882 results on '"Dipeptidases"'
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2. LOW RECOVERY OF CYSTINE FROM ENZYMIC HYDROLYSATES OF WHEAT FLOUR.
- Author
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TKACHUK R
- Subjects
- Chemical Phenomena, Chemistry, Cystine, Dipeptidases, Flour, Leucyl Aminopeptidase, Metabolism, Papain, Sulfur Isotopes, Thiazoles, Triticum
- Published
- 1964
- Full Text
- View/download PDF
3. [THE NATURE OF PEPTIDASES CONTAINED IN PREPARATIONS OF PH5-ENZYMES FROM RAT LIVER].
- Author
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SKARLAT IV, MOSOLOV VV, and AFANASEV PV
- Subjects
- Rats, Aminopeptidases, Chemical Phenomena, Chemistry, Dipeptidases, Glycine, Leucine, Leucyl Aminopeptidase, Liver enzymology, Peptide Hydrolases, Research
- Published
- 1964
4. Enzymatic determination of carnosine in meat and fish using β-Ala-Xaa dipeptidase and histidine ammonia-lyase derived from Pseudomonas putida NBRC100650
- Author
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Shinji Nagata, Hisashi Muramatsu, Tomoko Shimamura, Taisuke Harada, Daiki Sugihara, Karen Hashimoto, and Shin-ichiro Kato
- Subjects
chemistry.chemical_classification ,Dipeptidase ,Dipeptidases ,Meat ,biology ,Pseudomonas putida ,Carnosine ,Fishes ,biology.organism_classification ,Analytical Chemistry ,chemistry.chemical_compound ,Enzyme ,chemistry ,Biochemistry ,biology.protein ,Animals ,%22">Fish ,Cattle ,Histidine Ammonia-Lyase ,Histidine ammonia-lyase - Abstract
Carnosine is a naturally occurring dipeptide and a functional component in foods, while also showing health-promoting effects. Generally, food-derived carnosine is quantified via high-performance liquid chromatography (HPLC). We have developed a method for quantifying carnosine in foods using microbial enzymes, β-Ala-Xaa dipeptidase (BapA) and histidine ammonia-lyase (HAL). The carnosine concentrations in extracts of chicken, pork, beef, bonito, and tuna were determined via both HPLC and enzymatic determination. The carnosine contents measured via enzymatic determination were in agreement with those determined via conventional HPLC analysis. Relative standard-deviation values of the conventional HPLC method and the enzymatic determination of carnosine in foods were 0.728-5.76% and 0.504-4.58%, respectively. The recovery of carnosine in food extracts via enzymatic determination was 97-103%. Therefore, the developed enzymatic determination technique using BapA and HAL can be used for the determination of carnosine in meats and fishes with comparable accuracy to that of conventional HPLC analysis.
- Published
- 2022
5. Carnosine dipeptidase II (CNDP2) protects cells under cysteine insufficiency by hydrolyzing glutathione-related peptides
- Author
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Sho Kobayashi, Jia Han, Sohsuke Yamada, Junichi Fujii, Keita Nagaoka, Hideyo Sato, Nobuaki Okumura, Toshifumi Takao, Hiroyuki Konno, and Takujiro Homma
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Dipeptidases ,Kidney ,Carnosine ,Glutathione ,Oxidative phosphorylation ,Fibroblasts ,Biochemistry ,Embryonic stem cell ,Molecular biology ,Mice ,chemistry.chemical_compound ,Cytosol ,medicine.anatomical_structure ,chemistry ,Physiology (medical) ,Extracellular ,medicine ,Animals ,Macrophage ,Cysteine - Abstract
The knockout (KO) of the cystine transporter xCT causes ferroptosis, a type of iron-dependent necrotic cell death, in mouse embryonic fibroblasts, but this does not occur in macrophages. In this study, we explored the gene that supports cell survival under a xCT deficiency using a proteomics approach. Analysis of macrophage-derived peptides that were tagged with iTRAQ by liquid chromatography-mass spectrometry revealed a robust elevation in the levels of carnosine dipeptidase II (CNDP2) in xCT KO macrophages. The elevation in the CNDP2 protein levels was confirmed by immunoblot analyses and this elevation was accompanied by an increase in hydrolytic activity towards cysteinylglycine, the intermediate degradation product of glutathione after the removal of the γ-glutamyl group, in xCT KO macrophages. Supplementation of the cystine-free media of Hepa1-6 cells with glutathione or cysteinylglycine extended their survival, whereas the inclusion of bestatin, an inhibitor of CNDP2, counteracted the effects of these compounds. We established CNDP2 KO mice by means of the CRISPR/Cas9 system and found a decrease in dipeptidase activity in the liver, kidney, and brain. An acetaminophen overdose (350 mg/kg) showed not only aggravated hepatic damage but also renal injury in the CNDP2 KO mice, which was not evident in the wild-type mice that were receiving the same dose. The aggravated renal damage in the CNDP2 KO mice was consistent with the presence of abundant levels of CNDP2 in the kidney, the organ prone to developing ferroptosis. These collective data imply that cytosolic CNDP2, in conjugation with the removal of the γ-glutamyl group, recruits Cys from extracellular GSH and supports redox homeostasis of cells, particularly in epithelial cells of proximal tubules that are continuously exposed to oxidative insult from metabolic wastes that are produced in the body.
- Published
- 2021
6. Crystal structure of aspartyl dipeptidase from <scp> Xenopus laevis </scp> revealed ligand binding induced loop ordering and catalytic triad assembly
- Author
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Ashwani Kumar, Biplab Ghosh, Ravindra D. Makde, and R. Singh
- Subjects
Models, Molecular ,Dipeptidase ,Dipeptidases ,Protein Conformation ,Stereochemistry ,Xenopus ,Crystallography, X-Ray ,Ligands ,Biochemistry ,Xenopus laevis ,Enzyme activator ,chemistry.chemical_compound ,Structural Biology ,Catalytic Domain ,Catalytic triad ,Animals ,Amino Acid Sequence ,Molecular Biology ,chemistry.chemical_classification ,Aspartic Acid ,Binding Sites ,Dipeptide ,biology ,Active site ,Substrate (chemistry) ,biology.organism_classification ,Enzyme ,chemistry ,biology.protein - Abstract
Gene encoding aspartyl dipeptidase from Xenopus levies (PepExl) is upregulated by thyroid hormone and is proposed to play a significant role in resorption of tadpole tail during metamorphosis. However, the importance of peptidase activity for the resorption of the tail remain elusive. Here we report the crystal structures of first eukaryotic S51 peptidase, PepExl, in its ligand-free and Asp-bound states at 1.4 and 1.8 Å resolutions, respectively. The active site is located at dimeric interface and the catalytic triad is found to be dissembled in ligand-free and assembled in Asp-bound state. Structural comparison and molecular dynamic simulations of ligand-free and Asp-bound states shows that distinct loop (loop-A) plays an important role in active site shielding, substrate binding and enzyme activation. This study illuminates the Asp-X dipeptide binding in PepExl is associated with ordering of the loop-A and assembly of residues of catalytic triad in active conformation for enzymatic activity.
- Published
- 2021
7. Correlation between serum carnosinase concentration and renal damage in diabetic nephropathy patients
- Author
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Xiang-Ming Qi, Shiqi Zhang, Yong-Gui Wu, Benito A. Yard, Xue-Qi Liu, Zhou Zhou, and Qiu Zhang
- Subjects
0301 basic medicine ,Adult ,Male ,medicine.medical_specialty ,Dipeptidases ,Urinary system ,Clinical Biochemistry ,Serum albumin ,Renal function ,030209 endocrinology & metabolism ,Injury ,Diabetic nephropathy ,Carnosinase ,Kidney ,Biochemistry ,Gastroenterology ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Internal medicine ,medicine ,Albuminuria ,Humans ,Diabetic Nephropathies ,Cystatin C ,Creatinine ,biology ,business.industry ,Organic Chemistry ,Middle Aged ,medicine.disease ,030104 developmental biology ,chemistry ,Case-Control Studies ,biology.protein ,Disease Progression ,Kidney Failure, Chronic ,Microalbuminuria ,Original Article ,Female ,medicine.symptom ,business ,Biomarkers ,Glomerular Filtration Rate - Abstract
Diabetic nephropathy (DN) is one of the major complications of diabetes and contributes significantly towards end-stage renal disease. Previous studies have identified the gene encoding carnosinase (CN-1) as a predisposing factor for DN. Despite this fact, the relationship of the level of serum CN-1 and the progression of DN remains uninvestigated. Thus, the proposed study focused on clarifying the relationship among serum CN-1, indicators of renal function and tissue injury, and the progression of DN. A total of 14 patients with minimal changes disease (MCD) and 37 patients with DN were enrolled in the study. Additionally, 20 healthy volunteers were recruited as control. Further, DN patients were classified according to urinary albumin excretion rate into two groups: DN with microalbuminuria (n = 11) and DN with macroalbuminuria (n = 26). Clinical indicators including urinary protein components, serum carnosine concentration, serum CN-1 concentration and activity, and renal biopsy tissue injury indexes were included for analyzation. The serum CN-1 concentration and activity were observed to be the highest, but the serum carnosine concentration was the lowest in DN macroalbuminuria group. Moreover, within DN group, the concentration of serum CN-1 was positively correlated with uric acid (UA, r = 0.376, p = 0.026) and serum creatinine (SCr, r = 0.399, p = 0.018) and negatively correlated with serum albumin (Alb, r = − 0.348, p = 0.041) and estimated glomerular filtration rate (eGRF, r = − 0.432, p = 0.010). Furthermore, the concentration of serum CN-1 was discovered to be positively correlated with indicators including 24-h urinary protein–creatinine ratio (24 h-U-PRO/CRE, r = 0.528, p = 0.001), urinary albumin-to-creatinine ratio (Alb/CRE, r = 0.671, p = 0.000), urinary transferrin (TRF, r = 0.658, p = 0.000), retinol-binding protein (RBP, r = 0.523, p = 0.001), N-acetyl-glycosaminidase (NAG, r = 0.381, p = 0.024), immunoglobulin G (IgG, r = 0.522, p = 0.001), cystatin C (Cys-C, r = 0.539, p = 0.001), beta-2-microglobulin (β2-MG, r = 0.437, p = 0.009), and alpha-1-macroglobulin (α1-MG, r = 0.480, p = 0.004). Besides, in DN with macroalbuminuria group, serum CN-1 also showed a positive correlation with indicators of fibrosis, oxidative stress, and renal tubular injury. Taken together, our data suggested that the level of CN-1 was increased as clinical DN progressed. Thus, the level of serum CN-1 might be an important character during the occurrence and progression of DN. Our study will contribute significantly to future studies focused on dissecting the underlying mechanism of DN.
- Published
- 2021
8. Plasma prolidase levels are high in schizophrenia but not in first-episode psychosis
- Author
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Taner Öznur, Abdullah Bolu, Kamil Nahit Özmenler, Özcan Uzun, Onur Erdem, Sebla Ertuğrul, Mikail Burak Aydin, and S. Çetinkaya
- Subjects
Dipeptidases ,medicine.medical_specialty ,Psychosis ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Internal medicine ,First episode psychosis ,Healthy control ,polycyclic compounds ,medicine ,Humans ,heterocyclic compounds ,Pharmacology (medical) ,In patient ,Neurotransmitter ,Metalloproteinase ,business.industry ,organic chemicals ,medicine.disease ,030227 psychiatry ,carbohydrates (lipids) ,Psychiatry and Mental health ,Endocrinology ,Psychotic Disorders ,nervous system ,chemistry ,Schizophrenia ,Case-Control Studies ,business ,030217 neurology & neurosurgery - Abstract
An increasing number of studies have focussed on the neurobiology of schizophrenia (SCH), contributing to a better understanding of this disorder. Prolidase is a metalloprotease found in various tissues, which has been associated with the concentrations of proline, a neurotransmitter, in the brain. There is evidence to suggest that elevated proline levels play a role in SCH. The aim of the present study was to compare plasma proline levels in patients with drug-naive first-episode psychosis (FEP) and in those with SCH. Patients diagnosed with FEP (n = 26) and SCH (n = 26) were recruited for this study, in addition to healthy control volunteers (n = 26). Plasma prolidase levels were found to be elevated in the SCH group compared to drug-naive FEP and healthy control groups. This finding indicates that prolidase levels are higher in SCH patients, while levels in patients with drug-naive FEP are similar to those of healthy control. Follow-up studies are needed to provide a better understanding of prolidase in the etiopathogenesis of SCH.
- Published
- 2020
9. Human carnosinase 1 overexpression aggravates diabetes and renal impairment in BTBROb/Ob mice
- Author
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Benito A. Yard, Diego O. Pastene, Jiedong Qiu, Stefan Porubsky, Shiqi Zhang, Carolina Delatorre, Bernhard K. Krämer, Harry van Goor, Darya Nosan, Carsten Sticht, Xinmiao Zhang, Thomas Albrecht, Angelica Rodriguez-Niño, Sibylle J. Hauske, Groningen Institute for Organ Transplantation (GIOT), and Groningen Kidney Center (GKC)
- Subjects
Dipeptidases ,Gene Expression ,Mice, Obese ,Carnosine ,Diabetic nephropathy ,Kidney ,DISEASE ,Antioxidants ,PROTECTS ,Mice ,chemistry.chemical_compound ,0302 clinical medicine ,Drug Discovery ,Transgenic mice ,Diabetic Nephropathies ,Genetics (clinical) ,Proteinuria ,NOCTURNIN ,Immunohistochemistry ,medicine.anatomical_structure ,SECRETION ,Molecular Medicine ,Original Article ,medicine.symptom ,Glycosuria ,medicine.medical_specialty ,CNDP1 GENOTYPE ,NEPHROPATHY ,Mice, Transgenic ,METABOLISM ,Diabetes Mellitus, Experimental ,Nephropathy ,03 medical and health sciences ,Internal medicine ,medicine ,Animals ,Humans ,business.industry ,GLYCOSYLATION ,Gene Expression Profiling ,Computational Biology ,medicine.disease ,GENE ,Renal corpuscle ,Disease Models, Animal ,Endocrinology ,chemistry ,Glycated hemoglobin ,business ,Biomarkers ,030215 immunology - Abstract
Objective To assess the influence of serum carnosinase (CN1) on the course of diabetic kidney disease (DKD). Methods hCN1 transgenic (TG) mice were generated in a BTBROb/Ob genetic background to allow the spontaneous development of DKD in the presence of serum carnosinase. The influence of serum CN1 expression on obesity, hyperglycemia, and renal impairment was assessed. We also studied if aggravation of renal impairment in hCN1 TG BTBROb/Ob mice leads to changes in the renal transcriptome as compared with wild-type BTBROb/Ob mice. Results hCN1 was detected in the serum and urine of mice from two different hCN1 TG lines. The transgene was expressed in the liver but not in the kidney. High CN1 expression was associated with low plasma and renal carnosine concentrations, even after oral carnosine supplementation. Obese hCN1 transgenic BTBROb/Ob mice displayed significantly higher levels of glycated hemoglobin, glycosuria, proteinuria, and increased albumin-creatinine ratios (1104 ± 696 vs 492.1 ± 282.2 μg/mg) accompanied by an increased glomerular tuft area and renal corpuscle size. Gene-expression profiling of renal tissue disclosed hierarchical clustering between BTBROb/Wt, BTBROb/Ob, and hCN1 BTBROb/Ob mice. Along with aggravation of the DKD phenotype, 26 altered genes have been found in obese hCN1 transgenic mice; among them claudin-1, thrombospondin-1, nephronectin, and peroxisome proliferator–activated receptor-alpha have been reported to play essential roles in DKD. Conclusions Our data support a role for serum carnosinase 1 in the progression of DKD. Whether this is mainly attributed to the changes in renal carnosine concentrations warrants further studies. Key messages Increased carnosinase 1 (CN1) is associated with diabetic kidney disease (DKD). BTBROb/Ob mice with human CN1 develop a more aggravated DKD phenotype. Microarray revealed alterations by CN1 which are not altered by hyperglycemia. These genes have been described to play essential roles in DKD. Inhibiting CN1 could be beneficial in DKD.
- Published
- 2020
10. Secreted aspartyl peptidases by the emerging, opportunistic and multidrug-resistant fungal pathogens comprising the Candida haemulonii complex
- Author
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André L.S. Santos, Simone S.C. Oliveira, Marta H. Branquinha, Lys A. Braga-Silva, and Lívia S. Ramos
- Subjects
0106 biological sciences ,Dipeptidases ,Antifungal Agents ,Virulence ,Cathepsin D ,Biology ,01 natural sciences ,Virulence factor ,Microbiology ,03 medical and health sciences ,chemistry.chemical_compound ,Sequence Analysis, Protein ,Candida albicans ,Pepstatins ,Genetics ,Humans ,Protease Inhibitors ,Ecology, Evolution, Behavior and Systematics ,Candida ,030304 developmental biology ,0303 health sciences ,Candidiasis ,Aspartyl Peptidase ,Drug Resistance, Multiple ,Corpus albicans ,Multiple drug resistance ,Chemically defined medium ,Infectious Diseases ,chemistry ,Pepstatin ,010606 plant biology & botany - Abstract
The opportunistic pathogens comprising the Candida haemulonii complex (C. haemulonii, C. duobushaemulonii and C. haemulonii var. vulnera) are notable for their intrinsic resistance to different antifungal classes. Little is known about the virulence attributes in this emerging fungal complex. However, it is well-recognized that enzymes play important roles in virulence/pathogenesis of candidiasis. Herein, we aimed to identify aspartyl-type peptidases in 12 clinical isolates belonging to the C. haemulonii complex. All isolates were able to grow in a chemically defined medium containing albumin as the sole nitrogen source, and a considerable consumption of this protein occurred after 72–96 h. C. haemulonii var. vulnera isolates showed the lowest albumin degradation capability and the poorest growth rate. The measurement of secreted aspartyl peptidase (Sap) activity, using the cathepsin D fluorogenic substrate, varied from 91.6 to 413.3 arbitrary units and the classic aspartyl peptidase inhibitor, pepstatin A, significantly blocked the Sap released by C. haemulonii complex. No differences were observed in the Sap activity among the three fungal species. Flow cytometry, using a polyclonal antibody against Sap1-3 of C. albicans, detected homologous proteins at the surface of C. haemulonii complex (anti-Sap1-3-labeled cells ranged from 24.6 to 79.1%). Additionally, the immunoblotting assay, conducted with the same Sap1-3 antibody, recognized a protein of ∼50 kDa in all fungal isolates. A glimpse in the genome of these fungi revealed several potential proteins containing Sap1-3-like conserved domain. Altogether, our results demonstrated the potential of C. haemulonii species complex to produce Saps, an important virulence factor of Candida spp.
- Published
- 2020
11. Carnosine and Diabetic Nephropathy
- Author
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Verena Peters, Claus Peter Schmitt, and Benito A. Yard
- Subjects
0301 basic medicine ,Dipeptidases ,Antioxidant ,medicine.medical_treatment ,Carnosine ,Disease ,Type 2 diabetes ,Pharmacology ,Carbohydrate metabolism ,Kidney ,Biochemistry ,End stage renal disease ,Diabetic nephropathy ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Drug Discovery ,Humans ,Medicine ,Diabetic Nephropathies ,Major complication ,business.industry ,Organic Chemistry ,medicine.disease ,030104 developmental biology ,Diabetes Mellitus, Type 2 ,chemistry ,030220 oncology & carcinogenesis ,Molecular Medicine ,business - Abstract
Diabetic Nephropathy (DN) is a major complication in patients with type 1 or type 2 diabetes and represents the leading cause of end-stage renal disease. Novel therapeutic approaches are warranted. In view of a polymorphism in the carnosinase 1 gene CNDP1, resulting in reduced carnosine degradation activity and a significant DN risk reduction, carnosine (β-alanyl-L-histidine) has gained attention as a potential therapeutic target. Carnosine has anti-inflammatory, antioxidant, anti-glycation and reactive carbonyl quenching properties. In diabetic rodents, carnosine supplementation consistently improved renal histology and function and in most studies, also glucose metabolism. Even though plasma half-life of carnosine in humans is short, first intervention studies in (pre-) diabetic patients yielded promising results. The precise molecular mechanisms of carnosine mediated protective action, however, are still incompletely understood. This review highlights the recent knowledge on the role of the carnosine metabolism in DN.
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- 2020
12. Characterization of substrate specificity and novel autoprocessing mechanism of dipeptidase A from Prevotella intermedia
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Takayuki K. Nemoto, Mariko Naito, Takeshi Kobayakawa, Yuko Ohara-Nemoto, and Mohammad Tanvir Sarwar
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Models, Molecular ,0301 basic medicine ,Dipeptidase ,Dipeptidases ,Protein Conformation ,Clinical Biochemistry ,periodontal disease ,Locus (genetics) ,Prevotella intermedia ,Biochemistry ,Substrate Specificity ,law.invention ,Microbiology ,03 medical and health sciences ,0302 clinical medicine ,law ,dipeptidase A ,autoprocessing ,Molecular Biology ,chemistry.chemical_classification ,Lactobacillus helveticus ,biology ,Chemistry ,Streptococcus gordonii ,food and beverages ,030206 dentistry ,biology.organism_classification ,Amino acid ,030104 developmental biology ,biology.protein ,Recombinant DNA ,cysteine peptidase ,Cysteine - Abstract
Prevotella intermedia, a gram-negative anaerobic rod, is frequently observed in subgingival polymicrobial biofilm from adults with chronic periodontitis. Peptidases in periodontopathic bacteria are considered to function as etiological reagents. Pre. intermedia OMA14 cells abundantly express an unidentified cysteine peptidase specific for Arg-4-methycoumaryl-7- amide (MCA). BAU17746 (locus tag, PIOMA14_I_1238) and BAU18827 (locus tag, PIOMA14_II_0322) emerged as candidates of this peptidase from the substrate specificity and sequence similarity with C69-family Streptococcus gordonii Arg-aminopeptidase. The recombinant form of the former solely exhibited hydrolyzing activity toward Arg-MCA, and BAU17746 possesses a 26.6% amino acid identity with the C69-family Lactobacillus helveticus dipeptidase A. It was found that BAU17746 as well as L. helveticus dipeptidase A was a P1-position Arg-specific dipeptidase A, although the L. helveticus entity, a representative of the C69 family, had been reported to be specific for Leu and Phe. The fulllength form of BAU17746 was intramolecularly processed to a mature form carrying the N-terminus of Cys15. In conclusion, the marked Arg-MCA-hydrolyzing activity in Pre. intermedia was mediated by BAU17746 belonging to the C69-family dipeptidase A, in which the mature form carries an essential cysteine at the N-terminus., Biological Chemistry, 401(5), pp.629-642; 2020
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- 2020
13. Erythrocytes Prevent Degradation of Carnosine by Human Serum Carnosinase
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Henry, Oppermann, Stefanie, Elsel, Claudia, Birkemeyer, Jürgen, Meixensberger, and Frank, Gaunitz
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Serum ,reactive oxygen species ,Dipeptidases ,QH301-705.5 ,carnosinase ,Article ,carnosine ,Oxidative Stress ,Chemistry ,Adenosine Triphosphate ,erythrocytes ,liquid-chromatography mass spectrometry ,Humans ,Biology (General) ,QD1-999 - Abstract
The naturally occurring dipeptide carnosine (β-alanyl-l-histidine) has beneficial effects in different diseases. It is also frequently used as a food supplement to improve exercise performance and because of its anti-aging effects. Nevertheless, after oral ingestion, the dipeptide is not detectable in human serum because of rapid degradation by serum carnosinase. At the same time, intact carnosine is excreted in urine up to five hours after intake. Therefore, an unknown compartment protecting the dipeptide from degradation has long been hypothesized. Considering that erythrocytes may constitute this compartment, we investigated the uptake and intracellular amounts of carnosine in human erythrocytes cultivated in the presence of the dipeptide and human serum using liquid chromatography–mass spectrometry. In addition, we studied carnosine’s effect on ATP production in red blood cells and on their response to oxidative stress. Our experiments revealed uptake of carnosine into erythrocytes and protection from carnosinase degradation. In addition, no negative effect on ATP production or defense against oxidative stress was observed. In conclusion, our results for the first time demonstrate that erythrocytes can take up carnosine, and, most importantly, thereby prevent its degradation by human serum carnosinase.
- Published
- 2021
14. Aminopeptidases trim Xaa-Pro proteins, initiating their degradation by the Pro/N-degron pathway
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Alexander Varshavsky, Shun Jia Chen, Leehyeon Kim, and Hyun Kyu Song
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chemistry.chemical_classification ,Dipeptidases ,Saccharomyces cerevisiae Proteins ,Multidisciplinary ,biology ,Protein subunit ,Saccharomyces cerevisiae ,Biological Sciences ,biology.organism_classification ,Aminopeptidases ,Aminopeptidase ,Substrate Specificity ,Ubiquitin ligase ,Cytosol ,Enzyme ,chemistry ,Ubiquitin ,Biochemistry ,Proteolysis ,biology.protein ,Degron - Abstract
N-degron pathways are proteolytic systems that recognize proteins bearing N-terminal (Nt) degradation signals (degrons) called N-degrons. Our previous work identified Gid4 as a recognition component (N-recognin) of the Saccharomyces cerevisiae proteolytic system termed the proline (Pro)/N-degron pathway. Gid4 is a subunit of the oligomeric glucose-induced degradation (GID) ubiquitin ligase. Gid4 targets proteins through the binding to their Nt-Pro residue. Gid4 is also required for degradation of Nt-Xaa-Pro (Xaa is any amino acid residue) proteins such as Nt-[Ala-Pro]-Aro10 and Nt-[Ser-Pro]-Pck1, with Pro at position 2. Here, we show that specific aminopeptidases function as components of the Pro/N-degron pathway by removing Nt-Ala or Nt-Ser and yielding Nt-Pro, which can be recognized by Gid4-GID. Nt-Ala is removed by the previously uncharacterized aminopeptidase Fra1. The enzymatic activity of Fra1 is shown to be essential for the GID-dependent degradation of Nt-[Ala-Pro]-Aro10. Fra1 can also trim Nt-[Ala-Pro-Pro-Pro] (stopping immediately before the last Pro) and thereby can target for degradation a protein bearing this Nt sequence. Nt-Ser is removed largely by the mitochondrial/cytosolic/nuclear aminopeptidase Icp55. These advances are relevant to eukaryotes from fungi to animals and plants, as Fra1, Icp55, and the GID ubiquitin ligase are conserved in evolution. In addition to discovering the mechanism of targeting of Xaa-Pro proteins, these insights have also expanded the diversity of substrates of the Pro/N-degron pathway.
- Published
- 2021
15. Dipeptidase PEPDA Is Required for the Conidiation Pattern Shift in Metarhizium acridum
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Yuxian Xia, Xueling Su, Yueqing Cao, and Juan Li
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Hyphal growth ,Dipeptidase ,Dipeptidases ,Metarhizium ,Mutant ,Conidiation ,Applied Microbiology and Biotechnology ,Fungal Proteins ,Invertebrate Microbiology ,Amino Acids ,Gene ,Mycelium ,chemistry.chemical_classification ,Ecology ,biology ,Chemistry ,fungi ,Dipeptides ,Spores, Fungal ,biology.organism_classification ,Amino acid ,Biochemistry ,biology.protein ,Metarhizium acridum ,Food Science ,Biotechnology - Abstract
Filamentous fungi conduct two types of conidiation, typical conidiation from mycelia and microcycle conidiation (MC). Fungal conidiation can shift between the two patterns, which involves a large number of genes in the regulation of this process. In this study, we investigated the role of a dipeptidase gene pepdA in conidiation pattern shift in Metarhizium acridum, which is upregulated in MC pattern compared to typical conidiation. Results showed that disruption of the pepdA resulted in a shift of conidiation pattern from MC to typical conidiation. Metabolomic analyses of amino acids showed that the levels of 19 amino acids significantly changed in ΔpepdA mutant. The defect of MC in ΔpepdA can be rescued when nonpolar amino acids, α-alanine, β-alanine, or proline, were added into sucrose yeast extract agar (SYA) medium. Digital gene expression profiling analysis revealed that PEPDA mediated transcription of sets of genes which were involved in hyphal growth and development, sporulation, cell division, and amino acid metabolism. Our results demonstrated that PEPDA played important roles in the regulation of MC by manipulating the levels of amino acids in M. acridum. IMPORTANCE Conidia, as the asexual propagules in many fungi, are the start and end of the fungal life cycle. In entomopathogenic fungi, conidia are the infective form essential for their pathogenicity. Filamentous fungi conduct two types of conidiation, typical conidiation from mycelia and microcycle conidiation. The mechanisms of the shift between the two conidiation patterns remain to be elucidated. In this study, we demonstrated that the dipeptidase PEPDA, a key enzyme from the insect-pathogenic fungus Metarhizium acridum for the hydrolysis of dipeptides, is associated with a shift of conidiation pattern. The conidiation pattern of the ΔpepdA mutant was restored when supplemented with the nonpolar amino acids rather than polar amino acids. Therefore, this report highlights that the dipeptidase PEPDA regulates MC by manipulating the levels of amino acids in M. acridum.
- Published
- 2021
16. Role of prolidase activity and oxidative stress biomarkers in unexplained infertility
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Suzan Tabur, Sevsen Kulaksizoglu, Elif Isbilen, Özge Kömürcü, and Mahmut Kirmizioglu
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medicine.medical_specialty ,Dipeptidases ,medicine.medical_treatment ,medicine.disease_cause ,Internal medicine ,medicine ,Humans ,Patient group ,Unexplained infertility ,chemistry.chemical_classification ,Reactive oxygen species ,biology ,business.industry ,Vitamin E ,Obstetrics and Gynecology ,General Medicine ,Enzyme assay ,Antioxidant capacity ,Oxidative Stress ,Endocrinology ,chemistry ,Case-Control Studies ,Infertility ,biology.protein ,Positive relationship ,Female ,business ,Oxidative stress ,Biomarkers - Abstract
OBJECTIVE Our aim was to explore the significance of serum prolidase enzyme activity and oxidative stress in women with unexplained infertility (UEI). METHODS In this case-control study (n = 160; 86 cases; 74 controls) prolidase enzyme activity and total antioxidant status (TAS), total oxidant status (TOS), oxidative stress index (OSI), and vitamin E were measured in plasma using enzyme-linked immunosorbent assays. RESULTS Prolidase enzyme activity and TAS levels were particularly higher in the patient group (P = 0.013, P = 0.001, respectively). Decreased OSI levels were detected in the patient group (P = 0.001). There was a positive relationship of prolidase with vitamin E in both patient and control groups (r = 0.892, P = 0.001, and r = 0.659, P = 0.001, respectively). A positive, but weak, relationship was identified between prolidase activity and TOS levels and also between vitamin E and TOS levels in the UEI group (r = 0.265, P = 0.049, and r = 0.288, P = 0.014, respectively). No association was found between prolidase and TOS levels or between vitamin E and TOS levels in the control group (r = 0.0097, P = 0.527, and r = 0.085, P = 0.610, respectively). CONCLUSION Our results showed an association between serum prolidase activity and oxidative stress in UEI patients. Further studies including greater groups are required to show the role of reactive oxygen species in UEI.
- Published
- 2021
17. Novel Relationship Between Plasmalogen Lipid Signatures and Carnosine in Humans
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Peter J. Meikle, Negar Naderpoor, José Manuel Fernández-Real, Barbora de Courten, Jordi Mayneris-Perxachs, and Aya Mousa
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Adult ,Male ,medicine.medical_specialty ,Dipeptidases ,Lípids de la sang ,Plasmalogens ,Carnosine ,Adipokine ,Inflammation ,Carbohydrate metabolism ,chemistry.chemical_compound ,Young Adult ,Internal medicine ,Lipidomics ,medicine ,Glucose homeostasis ,Humans ,Obesity ,Muscle, Skeletal ,business.industry ,Muscles ,Músculs ,Cardiac muscle ,Interleukin-18 ,Lipidome ,Overweight ,Lipids ,medicine.anatomical_structure ,Endocrinology ,Glucose ,chemistry ,Blood lipids ,Obesitat ,Female ,medicine.symptom ,Insulin Resistance ,business ,Food Science ,Biotechnology - Abstract
Introduction Carnosine is a naturally occurring dipeptide abundant in the skeletal and cardiac muscle and brain, which has been shown to improve glucose metabolism and cardiovascular risk. This study showed that carnosine supplementation had positive changes on plasma lipidome. Here, this study aimed to establish the relationship of muscle carnosine and serum carnosinase-1 with cardiometabolic risk factors and the lipidome. Methods and Results This study profiles >450 lipid species in 65 overweight/obese nondiabetic individuals. Intensive metabolic testing is conducted using direct gold-standard measures of adiposity, insulin sensitivity and secretion, as well as measurement of serum inflammatory cytokines and adipokines. Muscle carnosine is negatively associated with 2-h glucose concentrations, whereas serum carnosinase-1 levels are negatively associated with insulin sensitivity and positively with IL-18. O-PLS and machine learning analyses reveal a strong association of muscle carnosine with ether lipids, particularly arachidonic acid-containing plasmalogens. Carnosinase-1 levels are positively associated with total phosphatidylethanolamines, but negatively with lysoalkylphosphatidylcholines, trihexosylceramides, and gangliosides. In particular, alkylphosphatidylethanolamine species containing arachidonic acid are positively associated with carnosinase-1. Conclusion These associations reinforce the role of muscle carnosine and serum carnosinase-1 in the interplay among low-grade chronic inflammation, glucose homeostasis, and insulin sensitivity This studyhas been funded by Instituto de Salud Carlos III through the project“PI18/01022” (Co-funded by European Regional Development Fund “Away to make Europe”). This work was supported by the National Healthand Medical Research Council (NHMRC) of Australia (APP1047897). JordiMayneris-Perxachs is funded by the Miguel Servet Program from the Insti-tuto de Salud Carlos III (ISCIII CP18/00009), co-funded by the EuropeanSocial Fund “Investing in your future.” A.M. is supported by a Peter Do-herty Biomedical Research Fellowship provided by the NHMRC. N.N. isa Monash Partners Health Services Research Fellow. B.dC. is supportedby a Royal Australasian College of Physicians Fellows Career DevelopmentFellowship. and is the recipient of the NHMRC grant, which funded thisstudy Open Access funding provided thanks to the CRUE-CSIC agreement with Wiley
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- 2021
18. DPP9’s Enzymatic Activity and Not Its Binding to CARD8 Inhibits Inflammasome Activation
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Sahana D. Rao, Cornelius Y. Taabazuing, Daniel A. Bachovchin, Andrew R. Griswold, Ashley J. Chui, Abir Bhattacharjee, and Daniel P. Ball
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0301 basic medicine ,Dipeptidases ,Inflammasomes ,Protein Conformation ,Mutant ,NLR Proteins ,Plasma protein binding ,01 natural sciences ,Biochemistry ,Organofluorophosphonates ,Serine ,03 medical and health sciences ,medicine ,Humans ,Protease Inhibitors ,Letters ,Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ,Adaptor Proteins, Signal Transducing ,chemistry.chemical_classification ,010405 organic chemistry ,HEK 293 cells ,Signal transducing adaptor protein ,Inflammasome ,General Medicine ,Neoplasm Proteins ,0104 chemical sciences ,Cell biology ,CARD Signaling Adaptor Proteins ,HEK293 Cells ,030104 developmental biology ,Enzyme ,chemistry ,Mutation ,Molecular Medicine ,Signal transduction ,Apoptosis Regulatory Proteins ,Protein Binding ,Signal Transduction ,medicine.drug - Abstract
Inflammasomes are multiprotein complexes formed in response to pathogens. NLRP1 and CARD8 are related proteins that form inflammasomes, but the pathogen-associated signal(s) and the molecular mechanisms controlling their activation have not been established. Inhibitors of the serine dipeptidyl peptidases DPP8 and DPP9 (DPP8/9) activate both NLRP1 and CARD8. Interestingly, DPP9 binds directly to NLRP1 and CARD8, and this interaction may contribute to the inhibition of NLRP1. Here, we use activity-based probes, reconstituted inflammasome assays, and mass spectrometry-based proteomics to further investigate the DPP9–CARD8 interaction. We show that the DPP9–CARD8 interaction, unlike the DPP9–NLRP1 interaction, is not disrupted by DPP9 inhibitors or CARD8 mutations that block autoproteolysis. Moreover, wild-type, but not catalytically inactive mutant, DPP9 rescues CARD8-mediated cell death in DPP9 knockout cells. Together, this work reveals that DPP9’s catalytic activity and not its binding to CARD8 restrains the CARD8 inflammasome and thus suggests the binding interaction likely serves some other biological purpose.
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- 2019
19. Quantitation of Methionine Sulfoxide in Milk and Milk-Based Beverages—Minimizing Artificial Oxidation by Anaerobic Enzymatic Hydrolysis
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Michael Hellwig and Martin Kölpin
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0106 biological sciences ,Dipeptidases ,food.ingredient ,Protein oxidation ,01 natural sciences ,Beverages ,Leucyl Aminopeptidase ,chemistry.chemical_compound ,Methionine ,food ,Tandem Mass Spectrometry ,Enzymatic hydrolysis ,Caffeic acid ,Animals ,Anaerobiosis ,Gallic acid ,food.beverage ,Chromatography ,Chemistry ,Evaporated milk ,Methionine sulfoxide ,Hydrolysis ,010401 analytical chemistry ,food and beverages ,General Chemistry ,0104 chemical sciences ,Coffee milk ,Milk ,Biocatalysis ,General Agricultural and Biological Sciences ,Oxidation-Reduction ,010606 plant biology & botany - Abstract
Protein oxidation in milk products may entail flavor changes through reactions at methionine residues. However, little is known about the extent of methionine oxidation in milk and milk products. In the present study, a method for quantitation of methionine, methionine sulfoxide, and methionine sulfone by a stable isotope dilution assay using HILIC-ESI-MS/MS was established. For the quantitation of protein-bound analytes, anaerobic enzymatic hydrolysis was optimized to suppress artificial methionine oxidation. Moreover, the method allowed for monitoring of artificial oxidation by coincubation of the labeled probe [2H8]methionine. The percentage of oxidized methionine was low in UHT milk (up to 1.6%) and evaporated milk (up to 8.8%), but higher in beverages such as cocoa milk drinks (up to 19.0%) and coffee milk drinks (up to 32.8%), resulting in methionine sulfoxide concentrations of up to 6.7 g/kg protein in the latter. These products are important dietary sources of methionine sulfoxide. Model studies revealed that methionine residues can be oxidized strongly in the presence of phenolic compounds such as catechin, caffeic acid, and gallic acid, which are present in cocoa and coffee and may account for the high extent of oxidation in commercial samples.
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- 2019
20. Platelet-Rich Plasma Promotes the Proliferation of Human Keratinocytes via a Progression of the Cell Cycle. A Role of Prolidase
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Jerzy Pałka, Ilona Oscilowska, Magdalena Misiura, Tomasz Guszczyn, Wojciech Miltyk, and Weronika Baszanowska
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0301 basic medicine ,Dipeptidases ,PRP ,PEPD ,lcsh:Chemistry ,0302 clinical medicine ,Cell Movement ,Epidermal growth factor receptor ,Receptor ,lcsh:QH301-705.5 ,Spectroscopy ,Skin ,biology ,integumentary system ,Platelet-Rich Plasma ,Chemistry ,Integrin beta1 ,General Medicine ,Cell cycle ,Computer Science Applications ,Cell biology ,prolidase ,ErbB Receptors ,030220 oncology & carcinogenesis ,cell cycle ,Cell Division ,Signal Transduction ,keratinocytes ,proliferation ,Article ,Catalysis ,Inorganic Chemistry ,03 medical and health sciences ,Humans ,Physical and Theoretical Chemistry ,Molecular Biology ,Cell Proliferation ,Wound Healing ,Cell growth ,HaCaT ,Organic Chemistry ,Fibroblasts ,030104 developmental biology ,Gene Expression Regulation ,lcsh:Biology (General) ,lcsh:QD1-999 ,Platelet-rich plasma ,biology.protein ,Wound healing - Abstract
Although the role of platelet-rich plasma (PRP) in tissue regeneration has been confirmed in many studies, the mechanism of this process is still not fully understood. Human keratinocytes (HaCaT) cells were used as an experimental model for studies on the effects of PRP on cell proliferation, migration, collagen biosynthesis, prolidase activity, and its expression and anabolic signaling. The activation of epidermal growth factor receptor (EGFR), &beta, 1-integrin, and insulin-like growth factor-1 receptor (IGF-1R) by PRP were investigated by western blot and immunocytochemistry. It has been found that PRP induced keratinocytes migration and proliferation through activation of cell cycle progression and EGFR downstream signaling. Similar biological effects were achieved by an addition to the culture medium of prolidase (PEPD), a ligand of EGFR (PRP is a rich source of PEPD&ndash, 2 ng/mL). PRP-dependent stimulation of collagen biosynthesis was accompanied by an increase in the expression of NF-&kappa, &beta, IGF-1R-downstream signaling proteins, and PEPD activity. The data suggest that PRP activates a complex of growth factors and adhesion receptors that stimulate cell proliferation, migration, and collagen biosynthesis. PRP induces PEPD-dependent human keratinocyte proliferation through activation of the EGFR receptor. Our study provides a novel mechanism of PRP-dependent wound healing.
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- 2021
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21. Extracellular Prolidase (PEPD) Induces Anabolic Processes through EGFR, β1-integrin, and IGF-1R Signaling Pathways in an Experimental Model of Wounded Fibroblasts
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Wojciech Miltyk, Jerzy Pałka, Weronika Baszanowska, Magdalena Misiura, and Ilona Oscilowska
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Dipeptidases ,medicine.medical_treatment ,EGFR ,PEPD ,wound healing ,Catalysis ,Article ,Receptor, IGF Type 1 ,lcsh:Chemistry ,Inorganic Chemistry ,Mice ,fibroblasts ,medicine ,Animals ,Humans ,Epidermal growth factor receptor ,Physical and Theoretical Chemistry ,Receptor ,β1-integrin ,lcsh:QH301-705.5 ,Molecular Biology ,Protein kinase B ,Spectroscopy ,PI3K/AKT/mTOR pathway ,Skin ,biology ,integumentary system ,Chemistry ,Growth factor ,Integrin beta1 ,Organic Chemistry ,General Medicine ,Computer Science Applications ,Cell biology ,prolidase ,ErbB Receptors ,lcsh:Biology (General) ,lcsh:QD1-999 ,Gene Expression Regulation ,biology.protein ,IGF-1 ,Signal transduction ,Wound healing ,Signal Transduction - Abstract
The role of prolidase (PEPD) as a ligand of the epidermal growth factor receptor (EGFR) was studied in an experimental model of wound healing in cultured fibroblasts. The cells were treated with PEPD (1&ndash, 100 nM) and analysis of cell viability, proliferation, migration, collagen biosynthesis, PEPD activity, and the expressions of EGFR, insulin-like growth factor 1 (IGF-1), and &beta, 1-integrin receptor including downstream signaling proteins were performed. It has been found that PEPD stimulated proliferation and migration of fibroblasts via activation of the EGFR-downstream PI3K/Akt/mTOR signaling pathway. Simultaneously, PEPD stimulated the expression of &beta, 1-integrin and IGF-1 receptors and proteins downstream to these receptors such as FAK, Grb2, and ERK1/2. Collagen biosynthesis was increased in control and &ldquo, wounded&rdquo, fibroblasts under PEPD treatment. The data suggest that PEPD-induced EGFR signaling may serve as a new attempt to therapy wound healing.
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- 2021
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22. Prolidase Stimulates Proliferation and Migration Through Activation of the PI3K/Akt/mTOR Signaling Pathway in Human Keratinocytes
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Jerzy Pałka, Magdalena Misiura, Ilona Ościłowska, Weronika Baszanowska, and Wojciech Miltyk
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0301 basic medicine ,Dipeptidases ,medicine.medical_treatment ,PEPD ,wound healing ,Ligands ,Receptor, IGF Type 1 ,lcsh:Chemistry ,Phosphatidylinositol 3-Kinases ,0302 clinical medicine ,Cell Movement ,Epidermal growth factor receptor ,lcsh:QH301-705.5 ,Spectroscopy ,biology ,Chemistry ,Integrin beta1 ,TOR Serine-Threonine Kinases ,General Medicine ,Computer Science Applications ,Cell biology ,prolidase ,ErbB Receptors ,030220 oncology & carcinogenesis ,Signal transduction ,Protein Binding ,Signal Transduction ,keratinocytes ,EGFR ,Models, Biological ,Article ,Catalysis ,Inorganic Chemistry ,03 medical and health sciences ,Cell Line, Tumor ,medicine ,Humans ,Physical and Theoretical Chemistry ,Molecular Biology ,Protein kinase B ,PI3K/AKT/mTOR pathway ,Cell Proliferation ,Cell growth ,Growth factor ,Organic Chemistry ,030104 developmental biology ,lcsh:Biology (General) ,lcsh:QD1-999 ,biology.protein ,Proto-Oncogene Proteins c-akt ,Transforming growth factor - Abstract
Recent reports have indicated prolidase (PEPD) as a ligand of the epidermal growth factor receptor (EGFR). Since this receptor is involved in the promotion of cell proliferation, growth, and migration, we aimed to investigate whether prolidase may participate in wound healing in vitro. All experiments were performed in prolidase-treated human keratinocytes assessing cell vitality, proliferation, and migration. The expression of downstream signaling proteins induced by EGFR, insulin-like growth factor 1 (IGF-1), transforming growth factor &beta, 1 (TGF-&beta, 1), and &beta, 1-integrin receptors were evaluated by Western immunoblotting and immunocytochemical staining. To determine collagen biosynthesis and prolidase activity radiometric and colorimetric methods were used, respectively. Proline content was determined by applying the liquid chromatography coupled with mass spectrometry. We found that prolidase promoted the proliferation and migration of keratinocytes through stimulation of EGFR-downstream signaling pathways in which the PI3K/Akt/mTOR axis was involved. Moreover, PEPD upregulated the expression of &beta, 1-integrin and IGF-1 receptors and their downstream proteins. Proline concentration and collagen biosynthesis were increased in HaCaT cells under prolidase treatment. Since extracellular prolidase as a ligand of EGFR induced cell growth, migration, and collagen biosynthesis in keratinocytes, it may represent a potential therapeutic approach for the treatment of skin wounds.
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- 2020
23. Serum prolidase, malondialdehyde and catalase levels for the evaluation of oxidative stress in patients with peripheral vertigo
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İsa Özbay, Muhammet Fatih Topuz, Cüneyt Kucur, Fatih Oghan, and Havva Kocak
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medicine.medical_specialty ,Dipeptidases ,Peripheral vertigo ,medicine.disease_cause ,Gastroenterology ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Internal medicine ,Malondialdehyde ,otorhinolaryngologic diseases ,medicine ,Humans ,In patient ,030223 otorhinolaryngology ,biology ,business.industry ,General Medicine ,Middle Aged ,Catalase ,Oxidative Stress ,Otorhinolaryngology ,chemistry ,030220 oncology & carcinogenesis ,Head and neck surgery ,biology.protein ,Vertigo ,Neurosurgery ,business ,Oxidative stress - Abstract
We aimed to evaluate oxidative stress in patients with peripheral vertigo by measuring serum prolidase, malondialdehyde (MDA) and catalase levels.A total of 30 patients (age: 60 ) with peripheral vertigo and 30 healthy subjects were recruited. Blood samples were collected from both groups and serum prolidase levels were measured using enzyme-linked immunosorbent assay (ELISA). MDA and catalase levels were measured by the spectrophotometric method.The most common cause of vertigo was BPPV (53.3%), followed by Ménière's disease (16.6%), vestibular neuritis (13.3%), lateral semicircular canal fistula (3.3%), and idiopathic vertigo (13.3%). Mean serum prolidase activity and MDA levels were significantly higher in the vertigo patients than in the control subjects (P 0.05); however, there was no statistically significant difference in mean serum catalase levels between the groups (P 0.05).We concluded that serum prolidase and MDA levels may be used as markers of oxidative stress in patients with peripheral vertigo.
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- 2020
24. Current Understanding of the Emerging Role of Prolidase in Cellular Metabolism
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Wojciech Miltyk and Magdalena Misiura
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Dipeptidases ,medicine.medical_treatment ,PEPD ,EGFR ,Review ,Catalysis ,lcsh:Chemistry ,Inorganic Chemistry ,Hydroxyproline ,chemistry.chemical_compound ,medicine ,Animals ,Humans ,Epidermal growth factor receptor ,Physical and Theoretical Chemistry ,Receptor ,lcsh:QH301-705.5 ,Molecular Biology ,Spectroscopy ,Cellular compartment ,Receptors, Interferon ,Prolidase deficiency ,biology ,Chemistry ,Growth factor ,Organic Chemistry ,General Medicine ,medicine.disease ,Computer Science Applications ,Cell biology ,prolidase ,ErbB Receptors ,lcsh:Biology (General) ,lcsh:QD1-999 ,biology.protein ,Tumor Suppressor Protein p53 ,cellular metabolism ,Transforming growth factor ,Signal Transduction - Abstract
Prolidase [EC 3.4.13.9], known as PEPD, cleaves di- and tripeptides containing carboxyl-terminal proline or hydroxyproline. For decades, prolidase has been thoroughly investigated, and several mechanisms regulating its activity are known, including the activation of the β1-integrin receptor, insulin-like growth factor 1 receptor (IGF-1) receptor, and transforming growth factor (TGF)-β1 receptor. This process may result in increased availability of proline in the mitochondrial proline cycle, thus making proline serve as a substrate for the resynthesis of collagen, an intracellular signaling molecule. However, as a ligand, PEPD can bind directly to the epidermal growth factor receptor (EGFR, epidermal growth factor receptor 2 (HER2)) and regulate cellular metabolism. Recent reports have indicated that PEPD protects p53 from uncontrolled p53 subcellular activation and its translocation between cellular compartments. PEPD also participates in the maturation of the interferon α/β receptor by regulating its expression. In addition to the biological effects, prolidase demonstrates clinical significance reflected in the disease known as prolidase deficiency. It is also known that prolidase activity is affected in collagen metabolism disorders, metabolic, and oncological conditions. In this article, we review the latest knowledge about prolidase and highlight its biological function, and thus provide an in-depth understanding of prolidase as a dipeptidase and protein regulating the function of key biomolecules in cellular metabolism.
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- 2020
25. Increased prolidase activity in Alzheimer's dementia: A case-control study
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Mathew Varghese, Ajit Bhalchandra Dahale, Thomas Gregor Issac, Geethu Krishna, Anu Kn Unni, Preeti Sinha, Palanimuthu T. Sivakumar, Shiva Shanker Reddy Mukku, Sarada Subramanian, and Lakshmi Prabha M
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medicine.medical_specialty ,Dipeptidases ,Extracellular matrix ,03 medical and health sciences ,0302 clinical medicine ,Alzheimer Disease ,Internal medicine ,medicine ,Humans ,Alzheimer s dementia ,Inverse correlation ,General Psychology ,chemistry.chemical_classification ,business.industry ,Case-control study ,General Medicine ,030227 psychiatry ,Extracellular Matrix ,Psychiatry and Mental health ,Enzyme ,Endocrinology ,chemistry ,Case-Control Studies ,Collagen metabolism ,business ,030217 neurology & neurosurgery - Abstract
Prolidase enzyme, which catalyzes the final step in collagen metabolism can influence the cognitive functions through changes in extracellular matrix (ECM) resulting in altered synaptic connectivity in Alzheimer's disease (AD). In this study, it was found that the prolidase activity was significantly higher (p = 0.0016) in AD subjects (5.62 ± 2.05 U/ mL) than control group (4.45 ± 0.92 U/ mL). The increase was significant beginning at mild AD (p = 0.006) with an inverse correlation with HMSE scores (p = 0.0344), thus implying that prolidase mediated alterations in ECM may be associated with the cognitive deficits seen in AD.
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- 2020
26. DPP8/9 inhibitors activate the CARD8 inflammasome in resting lymphocytes
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Daniel A. Bachovchin, Elizabeth L. Orth, Marian C. Okondo, Daniel P. Ball, Sahana D. Rao, Darren C. Johnson, and Hsin-Che Huang
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Cancer Research ,Programmed cell death ,Cell type ,Proteases ,Dipeptidases ,Inflammasomes ,Immunology ,NLR Proteins ,Lymphocyte Activation ,Article ,Cellular and Molecular Neuroscience ,Mice ,Cell death and immune response ,medicine ,Pyroptosis ,Animals ,Humans ,Protease Inhibitors ,Lymphocytes ,lcsh:QH573-671 ,Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ,Cells, Cultured ,Adaptor Proteins, Signal Transducing ,Innate immune system ,lcsh:Cytology ,Chemistry ,Immune cell death ,Cell Cycle ,Inflammasome ,Cell Biology ,Cell biology ,Neoplasm Proteins ,Rats ,CARD Signaling Adaptor Proteins ,Apoptosis Regulatory Proteins ,CD8 ,Intracellular ,medicine.drug - Abstract
Canonical inflammasomes are innate immune signaling platforms that are formed in response to intracellular pathogen-associated signals and trigger caspase-1-dependent pyroptosis. Inflammasome formation and signaling is thought to mainly occur in myeloid cells, and in particular monocytes and macrophages. Here we show that small molecule inhibitors of dipeptidyl peptidases 8 and 9 (DPP8/9), which activate the related CARD8 and NLRP1 inflammasomes, also activate pyroptosis in human and rodent resting lymphocytes. We found that both CD4+ and CD8+ T cells were particularly sensitive to these inhibitors, although the sensitivity of T cells, like macrophages, varied considerably between species. In human T cells, we show that CARD8 mediates DPP8/9 inhibitor-induced pyroptosis. Intriguingly, although activated human T cells express the key proteins known to be required for CARD8-mediated pyroptosis, these cells were completely resistant to DPP8/9 inhibitors. Overall, these data show that resting lymphoid cells can activate at least one inflammasome, revealing additional cell types and states poised to undergo rapid pyroptotic cell death in response to danger-associated signals.
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- 2020
27. Structure of human DPEP3 in complex with the SC-003 antibody Fab fragment reveals basis for lack of dipeptidase activity
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Vincent S. Stoll, Kristyn Hayashi, Sandro Vivona, Kenton L. Longenecker, Johannes Hampl, Aditi Prashar, and Patrick Koenig
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Dipeptidase ,Dipeptidases ,Immunoconjugates ,Pyrrolobenzodiazepine ,Epitope ,Antibodies ,03 medical and health sciences ,chemistry.chemical_compound ,Epitopes ,Immunoglobulin Fab Fragments ,Structural Biology ,Humans ,Asparagine ,Tyrosine ,Histidine ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,biology ,Chemistry ,030302 biochemistry & molecular biology ,Active site ,Membrane Proteins ,Enzyme ,Biochemistry ,Proteolysis ,biology.protein - Abstract
Dipeptidase 3 (DPEP3) is one of three glycosylphosphatidylinositol-anchored metallopeptidases potentially involved in the hydrolytic metabolism of dipeptides. While its exact biological function is not clear, DPEP3 expression is normally limited to testis, but can be elevated in ovarian cancer. Antibody drug conjugates targeting DPEP3 have shown efficacy in preclinical models with a pyrrolobenzodiazepine conjugate, SC-003, dosed in a phase I clinical trial (NCT02539719). Here we reveal the novel atomic structure of DPEP3 alone and in complex with the SC-003 Fab fragment at 1.8 and 2.8 A, respectively. The structure of DPEP3/SC-003 Fab complex reveals an eighteen-residue epitope across the DPEP3 dimerization interface distinct from the enzymatic active site. DPEP1 and DPEP3 extracellular domains share a conserved, dimeric TIM (β/α)8-barrel fold, consistent with 49% sequence identity. However, DPEP3 diverges from DPEP1 and DPEP2 in key positions of its active site: a histidine to tyrosine variation at position 269 reduces affinity for the β zinc and may cause substrate steric hindrance, whereas an aspartate to asparagine change at position 359 abolishes activation of the nucleophilic water/hydroxide, resulting in no in vitro activity against a variety of dipeptides and biological substrates (imipenem, leukotriene D4 and cystinyl-bis-glycine). Hence DPEP3, unlike DPEP1 and DPEP2, may require an activating co-factor in vivo or may remain an inactive, degenerate enzyme. This report sheds light on the structural discriminants between active and inactive membrane dipeptidases and provides a benchmark to characterize current and future DPEP3-targeted therapeutic approaches.
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- 2020
28. Co-expression with chaperones can affect protein 3D structure as exemplified by loss-of-function variants of human prolidase
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Manfred S. Weiss, Elżbieta Wątor, Piotr Wilk, and Maria Rutkiewicz
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Dipeptidases ,Mutant ,Biophysics ,Gene Expression ,Cleavage (embryo) ,Biochemistry ,03 medical and health sciences ,Protein Domains ,Structural Biology ,Loss of Function Mutation ,Heat shock protein ,Genetics ,medicine ,Humans ,Molecular Biology ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,Prolidase deficiency ,Chemistry ,C-terminus ,030302 biochemistry & molecular biology ,Cell Biology ,computer.file_format ,medicine.disease ,Protein Data Bank ,Recombinant Proteins ,Enzyme ,Inhouse research on structure dynamics and function of matter ,Protein folding ,computer ,Molecular Chaperones - Abstract
Prolidase catalyzes the cleavage of dipeptides containing proline on their C terminus. The reduction in prolidase activity is the cause of a rare disease named 'Prolidase Deficiency'. Local structural disorder was indicated as one of the causes for diminished prolidase activity. Previous studies showed that heat shock proteins can partially recover prolidase activity in vivo. To analyze this mechanism of enzymatic activity rescue, we compared the crystal structures of selected prolidase mutants expressed in the absence and in the presence of chaperones. Our results confirm that protein chaperones facilitate the formation of more ordered structures by their substrate protein. These results also suggest that the protein expression system needs to be considered as an important parameter in structural studies. DATABASES: The reported crystal structures and their associated structure factor amplitudes were deposited in the Protein Data Bank under the accession codes 6SRE, 6SRF, and 6SRG, respectively.
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- 2020
29. Decrease of the pro-inflammatory M1-like response by inhibition of dipeptidyl peptidases 8/9 in THP-1 macrophages quantitative proteomics of the proteome and secretome
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Aneta Stachowicz, Rafał Olszanecki, Anna Wiśniewska, Maciej Suski, Anna Kiepura, Józef Madej, Kamila Stachyra, Klaudia Czepiel, and Katarzyna Kuś
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Proteomics ,0301 basic medicine ,Dipeptidases ,Proteome ,THP-1 Cells ,Immunology ,Quantitative proteomics ,Macrophage polarization ,Context (language use) ,Models, Biological ,03 medical and health sciences ,0302 clinical medicine ,Immune system ,Humans ,Macrophage ,Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ,Molecular Biology ,Inflammation ,Chemistry ,Macrophages ,Endoplasmic reticulum ,Macrophage Activation ,Cell biology ,030104 developmental biology ,Apoptosis ,030215 immunology - Abstract
Background Cellular peptidases are an emerging target of novel pharmacological strategies in inflammatory diseases and cancer. In this context, the dipeptidyl peptidases 8 and 9 (DPP8/9) have gained special attention due to their activities in the immune cells. However, in spite of more than hundred protein substrates identified to date by mass spectrometry-based analysis, the cellular DPP8/9 functions are still elusive. Methods We applied the proteomic approach (iTRAQ-2DLC-MS/MS) to comprehensively analyze the role of DPP8/9 in the regulation of macrophage activation by in-depth protein quantitation of THP-1 proteome and secretome. Results Cells pre-incubated with DPP8/9 inhibitor (1G244) prior activation (LPS or IL-4/IL-13) diminished the expression levels of M1-like response markers, but not M2-like phenotype features. This was accompanied by multiple intra- and extra-cellular protein abundance changes in THP-1 cells, related to cellular metabolism, mitochondria and endoplasmic reticulum function, as well as those engaged with inflammatory and apoptotic processes, including previously reported and novel DPP8/9 targets. Conclusions Inhibition of DPP 8/9 had a profound effect on the THP-1 macrophage proteome and secretome, evidencing the decrease of the pro-inflammatory M1-like response. Presented results are to our best knowledge the first which, among others, highlight the metabolic effects of DPP8/9 inhibition in macrophages.
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- 2020
30. Overexpression of Prolidase Induces Autophagic Death in MCF-7 Breast Cancer Cells
- Author
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Adam Kazberuk, Agnieszka Klupczynska, Jerzy Pałka, Jan Matysiak, Joanna Teul, Arkadiusz Surażyński, Ilona Zaręba, and Thi Yen Ly Huynh
- Subjects
0301 basic medicine ,Dipeptidases ,Programmed cell death ,Proline ,Cell Survival ,Physiology ,Autophagic Cell Death ,Apoptosis ,Breast Neoplasms ,lcsh:Physiology ,lcsh:Biochemistry ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Proline dehydrogenase ,Biosynthesis ,Autophagy ,Proline Oxidase ,Humans ,lcsh:QD415-436 ,Viability assay ,Cells, Cultured ,lcsh:QP1-981 ,Proline oxidase ,Chemistry ,Fibroblasts ,Molecular biology ,030104 developmental biology ,Cell culture ,030220 oncology & carcinogenesis ,MCF-7 Cells ,Collagen - Abstract
Background/aims Proline availability for proline dehydrogenase/proline oxidase (PRODH/POX) may represent switching mechanism between PRODH/POX-dependent apoptosis and autophagy. The aim of the study was to evaluate the impact of overexpression of prolidase (proline releasing enzyme) on apoptosis/autophagy in breast cancer MCF-7 cells. Methods The model of MCF-7 cells with prolidase overexpression (MCF-7PL) was obtained. In order to targeting proline for PRODH/POX-dependent pathways substrate for prolidase, glycyl-proline (GP) was provided and proline utilization for collagen biosynthesis was blocked using 2-methoxyestradiol (MOE). Cell viability was determined using Nucleo-Counter NC-3000. The activity of prolidase was determined by colorimetric assay. DNA, collagen and total protein biosynthesis were determined by radiometric method. Expression of proteins was assessed by Western blot and immunofluorescence bioimaging. Concentration of proline was analyzed by liquid chromatography with mass spectrometry. Results Prolidase overexpression in MCF-7PL cells contributed to 10-fold increase in the enzyme activity, 3-fold increase in cytoplasmic proline level and decrease in cell viability and DNA biosynthesis compared to wild type MCF-7 cells. In MCF-7PL cells MOE and GP significantly decreased the number of living cells. MOE inhibited DNA biosynthesis in both cell lines while GP evoked inhibitory effect on the process only in MCF-7PL cells. In both cell lines, MOE or MOE+GP inhibited DNA and collagen biosynthesis. Although GP in MCF-7 cells stimulated collagen biosynthesis, it inhibited the process in MCF-7PL cells. The effects of studied compounds in MCF-7PL cells were accompanied by increase in the expression of Atg7, LC3A/B, Beclin-1, HIF-1α and decrease in the expression of PRODH/POX, active caspases-3 and -9. Conclusion The data suggest that overexpression of prolidase in MCF-7 cells contributes to increase in intracellular proline concentration and PRODH/POX-dependent autophagic cell death.
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- 2020
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31. A carnosine analog mitigates metabolic disorders of obesity by reducing carbonyl stress
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Giulio Vistoli, Lalage A. Katunga, Mara Colzani, Marina Carini, Luca Regazzoni, Stefania Gagliardi, Giuseppe Rossoni, Ettore Gilardoni, Danilo De Maddis, Giancarlo Aldini, Katsuhiko Funai, Ethan J. Anderson, Luca Cannizzaro, Renato Canevotti, and T. Blake Monroe
- Subjects
Male ,0301 basic medicine ,Dipeptidases ,Anserine ,Carnosine ,030209 endocrinology & metabolism ,macromolecular substances ,Pharmacology ,medicine.disease_cause ,Diabetes Mellitus, Experimental ,Rats, Sprague-Dawley ,Mice ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Insulin resistance ,Metabolic Diseases ,Diabetes mellitus ,medicine ,Animals ,Humans ,Obesity ,Muscle, Skeletal ,Aldehydes ,General Medicine ,medicine.disease ,Mice, Mutant Strains ,Rats ,Oxidative Stress ,030104 developmental biology ,chemistry ,Commentary ,Insulin Resistance ,Metabolic syndrome ,Steatohepatitis ,Oxidative stress ,Dyslipidemia ,Research Article - Abstract
Sugar- and lipid-derived aldehydes are reactive carbonyl species (RCS) frequently used as surrogate markers of oxidative stress in obesity. A pathogenic role for RCS in metabolic diseases of obesity remains controversial, however, partly because of their highly diffuse and broad reactivity and the lack of specific RCS-scavenging therapies. Naturally occurring histidine dipeptides (e.g., anserine and carnosine) show RCS reactivity, but their therapeutic potential in humans is limited by serum carnosinases. Here, we present the rational design, characterization, and pharmacological evaluation of carnosinol, i.e., (2S)-2-(3-amino propanoylamino)-3-(1H-imidazol-5-yl)propanol, a derivative of carnosine with high oral bioavailability that is resistant to carnosinases. Carnosinol displayed a suitable ADMET (absorption, distribution, metabolism, excretion, and toxicity) profile and was determined to have the greatest potency and selectivity toward α,β-unsaturated aldehydes (e.g., 4-hydroxynonenal, HNE, ACR) among all others reported thus far. In rodent models of diet-induced obesity and metabolic syndrome, carnosinol dose-dependently attenuated HNE adduct formation in liver and skeletal muscle, while simultaneously mitigating inflammation, dyslipidemia, insulin resistance, and steatohepatitis. These improvements in metabolic parameters with carnosinol were not due to changes in energy expenditure, physical activity, adiposity, or body weight. Collectively, our findings illustrate a pathogenic role for RCS in obesity-related metabolic disorders and provide validation for a promising new class of carbonyl-scavenging therapeutic compounds rationally derived from carnosine.
- Published
- 2018
32. Development of Low-Molecular Weight Collagen Peptide Complex with Glycosaminoglycan Components
- Author
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P. V. Shekhovtsov, T. I. Nikolaeva, V. V. Kaptsov, K. S. Laurinavichus, and M. V. Molchanov
- Subjects
0106 biological sciences ,Dipeptidases ,Swine ,Type II collagen ,Chymopapain ,Matrix (biology) ,Aminopeptidases ,01 natural sciences ,General Biochemistry, Genetics and Molecular Biology ,Glycosaminoglycan ,03 medical and health sciences ,Hydrolysis ,Chondrocytes ,0302 clinical medicine ,Enzymatic hydrolysis ,Papain ,medicine ,Animals ,Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ,Collagen Type II ,Glycosaminoglycans ,Molecular mass ,Hyaline cartilage ,Chemistry ,Cartilage ,Biological Transport ,General Medicine ,Molecular Weight ,Hyaline Cartilage ,medicine.anatomical_structure ,Biochemistry ,Cattle ,Muramidase ,Proteoglycans ,Peptides ,030217 neurology & neurosurgery ,010606 plant biology & botany - Abstract
Enzymatic hydrolysis of biopolymers of the cartilage tissue was studied for obtaining a complex of type II collagen peptides and glycosaminoglycan oligosaccharides. Hydrothermal hydrolysis in a high pressure homogenizer followed by enzymatic hydrolysis of the cartilage tissue biopolymers with proteolytic enzyme preparation Karipazim yielded a complex of collagen peptides and glycosaminoglycan oligosaccharides with molecular weights of 240-720 Da. Low molecular weight of the components increases their bioavailability. Entering into the cells (chondrocytes), low-molecular-weight peptides, disaccharides, and oligosaccharides as structural elements of the matrix can participate in the formation of fibrils of collagen and proteoglycans. Exogenous substances replenish deficient components of the matrix and/or their concentrations, affect the formation and strengthen the cartilage tissue. Thus, using cattle and porcine hyaline cartilages, we prepared a complex of biopolymers with lower molecular weights in comparison with previously developed nutraceuticals.
- Published
- 2018
33. DPP8 is a novel therapeutic target for multiple myeloma
- Author
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Ayumi Tatekoshi, Kotaro Arita, Paras Jawaid, Nam H. Dang, Noriaki Iwao, Takashi Kondo, Satoshi Iyama, Chikao Morimoto, Yusuke Kamihara, Miho Arai, Ryo Hatano, Mati Ur Rehman, Jun Murakami, Kohichi Takada, Akinori Wada, Kyo Noguchi, Tsutomu Sato, Kei Ohnuma, Ichiro Yasuda, and Sayaka Kajikawa
- Subjects
0301 basic medicine ,Programmed cell death ,Dipeptidases ,lcsh:Medicine ,Myeloma ,Apoptosis ,Antineoplastic Agents ,CD38 ,Article ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Mice, Inbred NOD ,Cell Line, Tumor ,Drug Discovery ,medicine ,Cytotoxic T cell ,Animals ,Humans ,Vildagliptin ,Protease Inhibitors ,lcsh:Science ,Multiple myeloma ,Multidisciplinary ,Chemistry ,lcsh:R ,Proteolytic enzymes ,medicine.disease ,030104 developmental biology ,Cell culture ,030220 oncology & carcinogenesis ,Cancer research ,lcsh:Q ,Female ,Multiple Myeloma ,medicine.drug - Abstract
Dipeptidyl peptidases (DPPs) are proteolytic enzymes that are ideal therapeutic targets in human diseases. Indeed, DPP4 inhibitors are widely used in clinical practice as anti-diabetic agents. In this paper, we show that DPP4 inhibitors also induced cell death in multiple human myeloma cells. Among five DPP4 inhibitors, only two of them, vildagliptin and saxagliptin, exhibited apparent cytotoxic effects on myeloma cell lines, without any difference in suppression of DPP4 activity. As these two DPP4 inhibitors are known to have off-target effects against DPP8/9, we employed the specific DPP8/9 inhibitor 1G244. 1G244 demonstrated anti-myeloma effects on several cell lines and CD138+ cells from patients as well as in murine xenograft model. Through siRNA silencing approach, we further confirmed that DPP8 but not DPP9 is a key molecule in inducing cell death induced by DPP8/9 inhibition. In fact, the expression of DPP8 in CD38+ cells from myeloma patients was higher than that of healthy volunteers. DPP8/9 inhibition induced apoptosis, as evidenced by activated form of PARP, caspases-3 and was suppressed by the pan-caspase inhibitor Z-VAD-FMK. Taken together, these results indicate that DPP8 is a novel therapeutic target for myeloma treatment.
- Published
- 2019
34. Structural basis for prolidase deficiency disease mechanisms
- Author
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Holger Dobbek, Uwe Mueller, Monika Uehlein, Piotr Wilk, Rafal Piwowarczyk, and Manfred S. Weiss
- Subjects
Models, Molecular ,0301 basic medicine ,Dipeptidases ,Proline ,Metallopeptidase ,Protein Conformation ,Protein Data Bank (RCSB PDB) ,Crystallography, X-Ray ,Biochemistry ,Substrate Specificity ,03 medical and health sciences ,Catalytic Domain ,medicine ,Humans ,Molecular Biology ,chemistry.chemical_classification ,Manganese ,Metalloproteinase ,Binding Sites ,Prolidase deficiency ,030102 biochemistry & molecular biology ,biology ,PEPD ,Active site ,Cell Biology ,medicine.disease ,Amino acid ,Hydroxyproline ,030104 developmental biology ,Enzyme ,chemistry ,Mutation ,Mutagenesis, Site-Directed ,biology.protein ,Mutant Proteins ,Prolidase Deficiency ,Protein Binding - Abstract
Prolidase is a metallopeptidase that cleaves iminodipeptides containing a proline (Pro) or hydroxyproline (Hyp) residue at their C-terminal end. The disease prolidase deficiency (PD) is a rare recessive human disorder characterized by reduced prolidase activity. PD manifests itself by a wide range of severe clinical symptoms, most commonly as skin ulceration, recurrent infections of the respiratory tract, and mental retardation. Several mutations in the PEPD gene have been identified that are responsible for the loss or the reduction of prolidase activity. In contrast, the structural basis of enzyme inactivation has so far remained elusive. In this study, we present high resolution crystal structures of a number of human prolidase (HsProl) variants, in which single amino acids are either substituted by others or deleted. The observed implications of the mutations on the three-dimensional structure of HsProl are reported and discussed and related to their enzymatic activity. The resulting structures may be divided into four groups depending on the presumed effect of the corresponding mutations on the reaction mechanism. The four possible inactivation mechanisms, which could be elucidated, are disruption of the catalytic Mn2 (OH- )-center, introduction of chain disorder along with the displacement of important active site residues, rigidification of the active site, and flexibilization of the active site. Database All refined structure coordinates as well as the corresponding structure factor amplitudes have been deposited in the PDB under the accession numbers 5MBY, 5MBZ, 5MC0, 5MC1, 5MC2, 5MC3, 5MC4, 5MC5, 6H2P, 6H2Q.
- Published
- 2018
35. Repurposing a bacterial prolidase for organophosphorus hydrolysis: Reshaped catalytic cavity switches substrate selectivity
- Author
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Jian Yang, Ru Li, Yu Liu, Yunzhu Xiao, and Lijuan Long
- Subjects
0106 biological sciences ,0301 basic medicine ,Dipeptidases ,Bioengineering ,Protein Engineering ,01 natural sciences ,Applied Microbiology and Biotechnology ,Chemical synthesis ,Paraoxon ,Catalysis ,Substrate Specificity ,03 medical and health sciences ,Hydrolysis ,Bacterial Proteins ,010608 biotechnology ,Catalytic Domain ,medicine ,Escherichia coli ,biology ,Chemistry ,Substrate (chemistry) ,Protein engineering ,Directed evolution ,Combinatorial chemistry ,030104 developmental biology ,biology.protein ,Enzyme promiscuity ,Directed Molecular Evolution ,Biotechnology ,medicine.drug - Abstract
Enzyme promiscuity is critical to the acquisition of evolutionary plasticity in cells and can be recruited for high-value chemical synthesis or xenobiotic degradation. The molecular determinants of substrate ambiguity are essential to this activity; however, these details remain unknown. Here, we performed the directed evolution of a prolidase to enhance its initially weak paraoxonase activity. The in vitro evolution led to an unexpected 1,000,000-fold switch in substrate selectivity, with a 30-fold increase in paraoxon hydrolysis and 40,000-fold decrease in peptide hydrolysis. Structural and in silico analyses revealed enlarged catalytic cavities and substrate repositioning as responsible for rapid catalytic transitions between distinct chemical reactions.
- Published
- 2019
36. Trichomonas vaginalis metalloproteinase TvMP50 is a monomeric Aminopeptidase P-like enzyme
- Author
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Enrique Rudiño-Piñera, Jonathan Puente-Rivera, María Elizbeth Alvarez-Sánchez, Rodrigo Arreola, Jorge Morales-Montor, and José Luis Villalpando
- Subjects
0301 basic medicine ,Dipeptidases ,Subfamily ,Protein Conformation ,Dimer ,Protozoan Proteins ,Bioengineering ,Crystallography, X-Ray ,Aminopeptidases ,Applied Microbiology and Biotechnology ,Biochemistry ,Aminopeptidase ,Substrate Specificity ,law.invention ,03 medical and health sciences ,chemistry.chemical_compound ,law ,Escherichia coli ,Trichomonas vaginalis ,Amino Acid Sequence ,Molecular Biology ,Phylogeny ,chemistry.chemical_classification ,Metalloproteinase ,Sequence Homology, Amino Acid ,Recombinant Proteins ,Molecular Weight ,030104 developmental biology ,Enzyme ,Monomer ,chemistry ,Metalloproteases ,Recombinant DNA ,Protein quaternary structure ,Biotechnology - Abstract
Previously, metalloproteinase was isolated and identified from Trichomonas vaginalis, belonging to the aminopeptidase P-like metalloproteinase subfamily A/B, family M24 of clan MG, named TvMP50. The native and recombinant TvMP50 showed proteolytic activity, determined by gelatin zymogram, and a 50 kDa band, suggesting that TvMP50 is a monomeric active enzyme. This was an unexpected finding since other Xaa-Pro aminopeptidases/prolidases are active as a biological unit formed by dimers/tetramers. In this study, the evolutionary history of TvMP50 and the preliminary crystal structure of the recombinant enzyme determined at 3.4 Å resolution is reported. TvMP50 was shown to be a type of putative, eukaryotic, monomeric aminopeptidase P, and the crystallographic coordinates showed a monomer on a "pseudo-homodimer" array on the asymmetric unit that resembles the quaternary structure of the M24B dimeric family and suggests a homodimeric aminopeptidase P-like enzyme as a likely ancestor. Interestingly, TvMP50 had a modified N-terminal region compared with other Xaa-Pro aminopeptidases/prolidases with three-dimensional structures; however, the formation of the standard dimer is structurally unstable in aqueous solution, and a comparably reduced number of hydrogen bridges and lack of saline bridges were found between subunits A/B, which could explain why TvMP50 portrays monomeric functionality. Additionally, we found that the Parabasalia group contains two protein lineages with a "pita bread" fold; the ancestral monomeric group 1 was probably derived from an ancestral dimeric aminopeptidase P-type enzyme, and group 2 has a probable dimeric kind of ancestral eukaryotic prolidase lineage. The implications of such hypotheses are also presented.
- Published
- 2018
37. Evaluation of serum prolidase activity and oxidative stress markers in men with BPH and prostate cancer
- Author
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Ahmet Celik, Faruk Kucukdurmaz, Sefa Resim, Hasan Dagli, Metin Kilinc, and E. Efe
- Subjects
Male ,0301 basic medicine ,Dipeptidases ,medicine.medical_specialty ,Prolidase ,Urology ,Prostatic Hyperplasia ,medicine.disease_cause ,urologic and male genital diseases ,lcsh:RC870-923 ,Superoxide dismutase ,03 medical and health sciences ,chemistry.chemical_compound ,Prostate cancer ,0302 clinical medicine ,medicine ,Humans ,Prospective Studies ,Aged ,Benign prostatic hyperplasia ,biology ,business.industry ,Prostatic Neoplasms ,Mean age ,General Medicine ,Middle Aged ,Hyperplasia ,Malondialdehyde ,medicine.disease ,lcsh:Diseases of the genitourinary system. Urology ,Ultrasound-Guided Prostate Biopsy ,Enzyme Activation ,030104 developmental biology ,Reproductive Medicine ,chemistry ,Oxidative stress ,030220 oncology & carcinogenesis ,Collagen metabolism ,biology.protein ,business ,Biomarkers ,Research Article - Abstract
Background Prostate cancer (PCa) and benign prostatic hyperplasia (BPH) are diseases of elderly men and are related to increased oxidative stress (OS). Although prolidase has a role in collagen metabolism, it is also used to evaluate OS in many diseases. However, there is a lack of data about serum prolidase activity (SPA) in prostate cancer. The aim of this study was to evaluate and compare SPA levels in males with BPH and PCa. Methods Evaluation was made of a total of 81 men who underwent transrectal ultrasound guided prostate biopsy for a definitive diagnosis due to high PSA levels. Patients were separated into 2 groups as BPH and PCa patients. Pre-biopsy malondialdehyde (MDA), superoxide dismutase (SOD), PSA levels and serum prolidase activities (SPA) were compared between the groups and the correlations of SPA with the other parameters were also investigated in both groups. Results BPH was diagnosed in 51 patients and PCa in 30. The mean age of patients was similar in both groups as 63.25 ± 5.81 years in the BPH group 65.30 ± 7.35 years in the PCa group(p:0.081). The median MDA and SOD levels were insignificantly increased in the PCa patients. SPA values were similar in BPH and PCa patients. SPA did not correlate with age, PSA, MDA or SOD levels in either group. Conclusions Our study results revealed that serum prolidase activity is similar in BPH and PCa cases and is not correlated with MDA, SOD or PSA levels.
- Published
- 2017
38. Anserine (beta-alanyl-3-methyl-L-histidine) improves neurovascular-unit dysfunction and spatial memory in aged AβPPswe/PSEN1dE9 Alzheimer’s-model mice
- Author
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Kota Enomoto, Jun Kaneko, Akiko Enya, Qiong Ding, and Tatsuhiro Hisatsune
- Subjects
0301 basic medicine ,medicine.medical_specialty ,Aging ,Dipeptidases ,Anserine ,Carnosine ,lcsh:Medicine ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,Mice ,0302 clinical medicine ,Alzheimer Disease ,Internal medicine ,medicine ,Presenilin-1 ,Elderly people ,Animals ,Humans ,Neutral ph ,lcsh:Science ,Episodic memory ,Spatial Memory ,Multidisciplinary ,Amyloid beta-Peptides ,Beta-Alanyl-3-Methyl-L-Histidine ,lcsh:R ,Brain ,Endothelial Cells ,Neurovascular bundle ,Disease Models, Animal ,030104 developmental biology ,Endocrinology ,Cerebral blood flow ,chemistry ,Immunology ,lcsh:Q ,Pericytes ,Neuroglia ,030217 neurology & neurosurgery - Abstract
Anserine/carnosine supplementation improves cerebral blood flow and verbal episodic memory in elderly people, as we previously reported. Anserine’s buffering activity is superior to that of carnosine at neutral pH. In human sera, carnosine but not anserine is rapidly cleaved by carnosinase, limiting its effectiveness. This study examined the effects of anserine on AβPPswe/PSEN1dE9 Alzheimer’s disease (AD) model mice over 18-months old, an age at which these mice exhibit detectable memory deficits. We found that 8 weeks of anserine treatment completely recovered the memory deficits, improved pericyte coverage on endothelial cells in the brain, and diminished chronic glial neuroinflammatory reactions in these mice. These results suggest that anserine (beta-alanyl-3-methyl-L-histidine) supplementation improved memory functions in AD-model mice by exerting a protective effect on the neurovascular units, which are composed of endothelial cells, pericytes, and supporting glial cells.
- Published
- 2017
39. Serum prolidase enzyme activity in obese subjects and its relationship with oxidative stress markers
- Author
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Ramazan Esen, Yasemin Usul Soyoral, Ufuk Düzenli, and Mehmet Aslan
- Subjects
Adult ,Male ,0301 basic medicine ,Dipeptidases ,medicine.medical_specialty ,Antioxidant ,medicine.medical_treatment ,Clinical Biochemistry ,Connective tissue ,030204 cardiovascular system & hematology ,medicine.disease_cause ,Biochemistry ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,medicine ,Humans ,Obesity ,biology ,Chemistry ,Biochemistry (medical) ,General Medicine ,medicine.disease ,Enzyme assay ,Oxidative Stress ,030104 developmental biology ,medicine.anatomical_structure ,Endocrinology ,Case-Control Studies ,biology.protein ,Population study ,Female ,Obese subjects ,Body mass index ,Biomarkers ,Oxidative stress - Abstract
The relationship between increased serum enzyme activity of prolidase and increased rate of collagen turnover in the arterial wall has been asserted in previous studies. Collagen reflects much of the strength to the connective tissue involved in the arterial wall. Atherosclerosis is very common vessel disease and oxidative stress plays a pivotal role in the etiopathogenesis. Our objective was to examine the serum enzyme activity of prolidase and its possible relationships with oxidative stress parameters in obese subjects.Our present study was conducted 27 obese subjects and 26 age-matched healthy control subjects. The serum enzyme activity of prolidase in all study population was evaluated spectrophotometrically. Oxidative stress levels in obese subjects were analyzed with total antioxidant capacity (TAC) and total oxidant status (TOS) as well as oxidative stress index (OSI).Obese subjects have higher serum TOS and OSI indicators as well as prolidase activity than those in control subjects (for all; p0.001). Moreover, obese subjects have lower levels of TAC than in those in healthy subjects (p0.001). In the Pearson's correlation analysis, enzyme activity of prolidase was positively related with TOS (p0.001, r=0.529) and OSI (p0.001, r=0.519) as well as BMI (p0.001, r=0.692) and inversely related with TAC (p0.05, r=-0.405) in obese subjects.Increased serum prolidase activity and decreased antioxidant levels are likely to be a results of increased of oxidative stress levels in obese subjects. The significantly correlation between increased oxidative stress and increased prolidase activity may play a pivotal role in etiopathogenesis of atherosclerotic cardiovascular diseases in obese subjects.
- Published
- 2017
40. Crystallographic structure of recombinant Lactococcus lactis prolidase to support proposed structure-function relationships
- Author
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Oarabile Kgosisejo, Jian An Chen, Pawel Grochulski, and Takuji Tanaka
- Subjects
0301 basic medicine ,Dipeptidases ,Protein Conformation ,Stereochemistry ,Dimer ,Allosteric regulation ,Biophysics ,Crystallography, X-Ray ,Biochemistry ,Substrate Specificity ,Analytical Chemistry ,Structure-Activity Relationship ,03 medical and health sciences ,chemistry.chemical_compound ,Pyrococcus horikoshii ,Pyrococcus ,Allosteric Regulation ,Catalytic Domain ,Amino Acid Sequence ,Molecular Biology ,Peptide sequence ,chemistry.chemical_classification ,Binding Sites ,030102 biochemistry & molecular biology ,biology ,Lactococcus lactis ,Active site ,biology.organism_classification ,Amino acid ,Crystallography ,030104 developmental biology ,chemistry ,Mutagenesis, Site-Directed ,biology.protein ,Protein Multimerization - Abstract
Lactococcus lactis prolidase (Xaa-Pro dipeptidase; EC.3.4.13.9) is unique from other prolidases by showing allosteric behaviour, substrate inhibition, and metal-dependent substrate specificity. We have previously shown several critical residues for these characteristics using site-directed mutagenesis and amino acid sequence-based models. In this present study, the three-dimensional structure of recombinant L. lactis prolidase was determined by X-ray crystallography at 2.25Å resolution in order to provide evidences of the proposed mechanism. Three molecules are located in the crystal asymmetric unit where molecule A forms a dimer with molecule B, while molecule C forms a dimer with molecule C' in the adjacent asymmetric unit. Of all the three molecules, molecule C is less defined and incomplete. While this fact compromises the overall quality of the refined model, the functional interpretation of the structure is not compromised since the biologically-functional homodimeric configuration of L. lactis prolidase is represented by well-defined molecules A and B. The refined model confirmed that there is a twelve-residue (residues 32-43) loop structure from one subunit over the active site of the other subunit, proving the existence of the putative loop structure in our previous study. This loop is three amino acids longer than the loops of prolidases of Pyrococcus furious (1pv9) and Pyrococcus horikoshii OT3 (1wy2). The crystal structure shows the loop structure can form two states ("open" and "closed" states) through interaction between the loop and active site proximity. It supports the proposed formation of allosteric site by the loop and Arg 293.
- Published
- 2017
41. Carbon nanotube as a carrier in drug delivery system for carnosine dipeptide: A computer simulation study
- Author
-
Leila Rahmani and Sepideh Ketabi
- Subjects
Dipeptidases ,Carnosine ,Bioengineering ,02 engineering and technology ,Carbon nanotube ,010402 general chemistry ,01 natural sciences ,law.invention ,Biomaterials ,chemistry.chemical_compound ,Drug Delivery Systems ,law ,Organic chemistry ,Computer Simulation ,Solubility ,Drug Carriers ,Aqueous solution ,Nanotubes, Carbon ,Solvation ,Interaction energy ,021001 nanoscience & nanotechnology ,0104 chemical sciences ,Solutions ,Chemical engineering ,chemistry ,Mechanics of Materials ,Zwitterion ,Solvents ,Quantum Theory ,Thermodynamics ,Density functional theory ,0210 nano-technology ,Monte Carlo Method - Abstract
Biological application of carbon nanotube in drug delivery is our main concern in this investigation. For this purpose interaction of carnosine and carbon nanotube was studied in both gas phase and separately in aqueous media. Three possible interactions of carnosine dipeptide with (5,5) carbon nanotube in physiological media were considered. At first step each species were modeled using quantum mechanical calculations, in the next step, their properties in aqueous solution were studied by applying Monte Carlo simulations. The results of density functional calculations in gas phase showed that interaction of zwitterion of carnosine with carbon nanotube via NH3+ had relatively higher interaction energy than the other complexes. Computation of solvation free energies in water showed functionalization with carnosine enhanced the solubility of carbon nanotube significantly that improve the medicinal applications of these materials. Calculation of complexation free energies indicated that zwitterion of carnosine with carbon nanotube via NH3+ produced the most stable complex in aqueous solution. This tendency could be observed in gas and liquid phase similarly.
- Published
- 2017
42. Allosteric inhibition of carnosinase (CN1) by inducing a conformational shift
- Author
-
Christian Thiel, Thomas Fleming, Markus Hecker, Andreas H. Wagner, Vittorio Calabrese, Verena Peters, Peter P. Nawroth, Claus Peter Schmitt, Elisabete Forsberg, Tim Weigand, Antje van den Berg, Giulio Vistoli, and Kristina Klingbeil
- Subjects
Carnosinase 1 activity ,0301 basic medicine ,Dipeptidase ,Dipeptidases ,Allosteric regulation ,Molecular Conformation ,Carnosine ,Molecular Dynamics Simulation ,allosteric inhibition ,law.invention ,Diabetic nephropathy ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Allosteric Regulation ,law ,Drug Discovery ,medicine ,Humans ,CN1 ,Ammonium ,Sulfhydryl Compounds ,Enzyme Inhibitors ,glutathione ,Pharmacology ,diabetes ,biology ,Drug Discovery3003 Pharmaceutical Science ,N-acetylcysteine ,lcsh:RM1-950 ,General Medicine ,Glutathione ,medicine.disease ,lcsh:Therapeutics. Pharmacology ,030104 developmental biology ,chemistry ,Biochemistry ,030220 oncology & carcinogenesis ,Recombinant DNA ,biology.protein ,Research Paper ,Cysteine - Abstract
In humans, low serum carnosinase (CN1) activity protects patients with type 2 diabetes from diabetic nephropathy. We now characterized the interaction of thiol-containing compounds with CN1 cysteine residue at position 102, which is important for CN1 activity. Reduced glutathione (GSH), N-acetylcysteine and cysteine (3.2 ± 0.4, 2.0 ± 0.3, 1.6 ± 0.2 µmol/mg/h/mM; p
- Published
- 2017
43. The Effects of Hyperbaric Oxygen Treatment on Total Antioxidant Capacity and Prolidase Activity after Bile Duct Ligation in Rats
- Author
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Pinar Atukeren, Hayriye Erman, Burhan Aksu, Duygu Terzioglu, Lebriz Uslu, Suleyman Ayvaz, Gonul Simsek, Hafize Uzun, and Remise Gelisgen
- Subjects
Male ,Dipeptidases ,medicine.medical_specialty ,Antioxidant ,medicine.medical_treatment ,medicine.disease_cause ,digestive system ,Antioxidants ,Rats, Sprague-Dawley ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Cholestasis ,Predictive Value of Tests ,Fibrosis ,Internal medicine ,Animals ,Humans ,Medicine ,Ligation ,Common Bile Duct ,chemistry.chemical_classification ,Hyperbaric Oxygenation ,business.industry ,Bile duct ligation ,medicine.disease ,N-Acetylneuraminic Acid ,Rats ,Sialic acid ,Oxygen ,Disease Models, Animal ,Oxidative Stress ,Antioxidant capacity ,Endocrinology ,Liver ,chemistry ,Biochemistry ,Spectrophotometry ,030220 oncology & carcinogenesis ,Thiol ,030211 gastroenterology & hepatology ,Surgery ,business ,Biomarkers ,Oxidative stress - Abstract
BACKGROUND Hyperbaric oxygen (HBO) therapy may improve cholestasis, increase hepatic regeneration, and decrease oxidative stress in liver. In this study, we aimed to investigate the effects of HBO therapy on hepatic oxidative stress parameters, such as total thiol groups (T-SH), protein carbonyl (PCO), and total antioxidant capacity (TAC) as well as the predictive value of the noninvasive biochemical marker, sialic acid (SA), and prolidase activity in bile duct ligation (BDL)-induced oxidative damage and fibrosis in rats. METHODS We divided 32 adult male Sprague Dawley rats into four groups: sham, sham + HBO, BDL, and BDL + HBO; each group contained eight animals. We placed the sham + HBO and BDL + HBO groups in an experimental hyperbaric chamber, in which we administered pure oxygen at 2.5 atmospheres for 90 min on 14 consecutive days. RESULTS The application of BDL significantly increased PCO levels and prolidase activity, and decreased T-SH and TAC levels. HBO significantly decreased PCO levels and prolidase activity and increased T-SH and TAC levels in the liver tissues. There was no significant difference in sialic acid levels between any groups. CONCLUSIONS These results indicate that HBO therapy has hepatoprotective effects on BDL-induced injury by decreasing PCO and prolidase activity and increasing antioxidant activities. We therefore suggest that HBO therapy may be useful after BDL-induced injury.
- Published
- 2016
44. DPP8 and DPP9 inhibition induces pro-caspase-1-dependent monocyte and macrophage pyroptosis
- Author
-
Sarah E. Poplawski, David G. Sanford, Mitchell S. Wang, Eun Bin Go, Daniel A. Bachovchin, William W. Bachovchin, Yuxin Liu, Darren C. Johnson, Michael O Arciprete, Ramya Sridharan, Ashley J. Chui, Wengen Wu, Marian C. Okondo, Todd R. Golub, and Jack H. Lai
- Subjects
0301 basic medicine ,Dipeptidases ,Proteases ,Programmed cell death ,Serine Proteinase Inhibitors ,Molecular Conformation ,Caspase 1 ,Article ,Cell Line ,Serine ,Mice ,Structure-Activity Relationship ,03 medical and health sciences ,Pyroptosis ,medicine ,Animals ,Humans ,Macrophage ,Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ,Molecular Biology ,Dose-Response Relationship, Drug ,Chemistry ,Macrophages ,Monocyte ,Dipeptides ,Cell Biology ,Boronic Acids ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,Lytic cycle ,Leukocytes, Mononuclear - Abstract
Val-boroPro (Talabostat, PT-100), a nonselective inhibitor of post-proline cleaving serine proteases, stimulates mammalian immune systems through an unknown mechanism of action. Despite this lack of mechanistic understanding, Val-boroPro has attracted substantial interest as a potential anticancer agent, reaching phase 3 trials in humans. Here we show that Val-boroPro stimulates the immune system by triggering a proinflammatory form of cell death in monocytes and macrophages known as pyroptosis. We demonstrate that the inhibition of two serine proteases, DPP8 and DPP9, activates the pro-protein form of caspase-1 independent of the inflammasome adaptor ASC. Activated pro-caspase-1 does not efficiently process itself or IL-1β but does cleave and activate gasdermin D to induce pyroptosis. Mice lacking caspase-1 do not show immune stimulation after treatment with Val-boroPro. Our data identify what is to our knowledge the first small molecule that induces pyroptosis and reveals a new checkpoint that controls the activation of the innate immune system.
- Published
- 2016
45. Family-wide annotation of enzymatic pathways by parallel in vivo metabolomics
- Author
-
Veronica L. Li, Jonathan Z. Long, Joon T. Kim, Stephanie M. Terrell, and Curt R. Fischer
- Subjects
Male ,Dipeptidases ,Spectrometry, Mass, Electrospray Ionization ,Clinical Biochemistry ,Computational biology ,Biology ,01 natural sciences ,Biochemistry ,Article ,Amidohydrolases ,chemistry.chemical_compound ,Mice ,Metabolomics ,In vivo ,Drug Discovery ,Animals ,Humans ,Molecular Biology ,Chromatography, High Pressure Liquid ,Pharmacology ,chemistry.chemical_classification ,Mice, Knockout ,Dipeptide ,010405 organic chemistry ,Catabolism ,Hydrolysis ,Dipeptides ,Recombinant Proteins ,0104 chemical sciences ,Up-Regulation ,Mice, Inbred C57BL ,Metabolic pathway ,Enzyme ,HEK293 Cells ,chemistry ,Liver ,Mutagenesis, Site-Directed ,Molecular Medicine ,ACY1 ,Homeostasis - Abstract
Enzymes catalyze fundamental biochemical reactions that control cellular and organismal homeostasis. Here we present an approach for de novo biochemical pathway discovery across entire enzyme families using parallel viral transduction in mice and untargeted liquid chromatography-mass spectrometry. Applying this method to the mammalian M20 peptidases uncovers both known pathways of amino acid metabolism as well as a previously unknown CNDP2-regulated pathway for threonyl dipeptide catabolism. Ablation of CNDP2 in mice elevates threonyl dipeptides across multiple tissues, establishing the physiologic relevance of our biochemical assignments. Taken together, these data underscore the utility of parallel in vivo metabolomics for the family-wide discovery of enzymatic pathways. Enzymes catalyze fundamental biochemical reactions that control cellular and organismal homeostasis. Here we present an approach for de novo biochemical pathway discovery across entire enzyme families using parallel viral transduction in mice and untargeted liquid chromatography-mass spectrometry. Applying this method to the mammalian M20 peptidases uncovers known pathways of amino acid metabolism mediated by ACY1 (hydrolysis of N-acetyl amino acids) and CNDP2 (hydrolysis of carnosine). We also uncover a previously unknown CNDP2-regulated pathway for threonyl dipeptide catabolism. Ablation of CNDP2 in mice elevates threonyl dipeptides across multiple tissues, establishing the physiologic relevance of our biochemical assignments. Taken together, these data underscore the utility of parallel in vivo metabolomics for the family-wide discovery of enzymatic pathways.
- Published
- 2019
46. CNDP2 Acts as an Activator for Human Ovarian Cancer Growth and Metastasis via the PI3K/AKT Pathway
- Author
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Ling P Zhao, Li Q Zhang, Hong S Lu, Hua Q Yang, Xian J Chen, Su Q Yang, and Ying Wang
- Subjects
Cancer Research ,Dipeptidases ,endocrine system diseases ,growth ,medicine.disease_cause ,Metastasis ,03 medical and health sciences ,Mice ,Phosphatidylinositol 3-Kinases ,0302 clinical medicine ,Cell Movement ,Cell Line, Tumor ,medicine ,Animals ,Humans ,metastasis ,Neoplasm Metastasis ,PI3K/AKT/mTOR pathway ,030304 developmental biology ,Cell Proliferation ,Neoplasm Staging ,Mice, Knockout ,Ovarian Neoplasms ,0303 health sciences ,Chemistry ,Activator (genetics) ,medicine.disease ,Xenograft Model Antitumor Assays ,Gene Expression Regulation, Neoplastic ,CNDP2 ,Cytosol ,Disease Models, Animal ,Cell Transformation, Neoplastic ,ovarian cancer ,Oncology ,030220 oncology & carcinogenesis ,Molecular mechanism ,Cancer research ,Female ,Original Article ,Ovarian cancer ,Carcinogenesis ,Proto-Oncogene Proteins c-akt ,Signal Transduction - Abstract
Introduction: The mechanism of tumorigenesis and metastasis of ovarian cancer has not yet been elucidated. This study aimed to investigate the role and molecular mechanism of cytosolic nonspecific dipeptidase 2 in tumorigenesis and metastasis. Methods: Cytosolic nonspecific dipeptidase 2 expression in human ovarian cancer tissues and cell lines was assessed with methyl thiazolyl tetrazolium (MTT), clone formation, and transwell assays performed to evaluate the ability of ovarian cancer cells to proliferate and migrate. Nude mice tumor formation experiments were also performed by subcutaneously injecting cells with stable cytosolic nonspecific dipeptidase 2 knockdown and control SKOV3 cells into BALB/c female nude mice to detect changes in PI3K/AKT pathway-related proteins by Western blotting. Results: Cytosolic nonspecific dipeptidase 2 was highly expressed in human ovarian cancer tissues, with its expression associated with pathological data, including ovarian cancer metastasis. A cytosolic nonspecific dipeptidase 2 stable knockdown or ectopic expression ovarian cancer cell model was established and demonstrated that cytosolic nonspecific dipeptidase 2 could promote the proliferation of ovarian cancer cells. Transwell cell migration and invasion assays confirmed that cytosolic nonspecific dipeptidase 2 enhanced cell metastasis in ovarian cancer. Furthermore, in vivo xenograft experiments demonstrated that cytosolic nonspecific dipeptidase 2 can promote the development and progression of ovarian cancer, increasing the expression of phosphorylated PI3K and AKT. Conclusions: Cytosolic nonspecific dipeptidase 2 promotes the occurrence and development of ovarian cancer through the PI3K/AKT signaling pathway.
- Published
- 2019
47. Engineering Xaa-Pro dipeptidyl aminopeptidase for specific cleavage of glucagon and glucagon-like peptide 1 from fusion proteins
- Author
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Allan Christian Shaw, Erik Vernet, Marie Østergaard Pedersen, and Henning Thøgersen
- Subjects
0106 biological sciences ,Proteases ,Dipeptidases ,medicine.medical_treatment ,Genetic Vectors ,Gene Expression ,Peptide ,Protein Engineering ,01 natural sciences ,Aminopeptidase ,03 medical and health sciences ,chemistry.chemical_compound ,Bacterial Proteins ,Glucagon-Like Peptide 1 ,010608 biotechnology ,medicine ,Escherichia coli ,Humans ,Cloning, Molecular ,030304 developmental biology ,Enzyme Assays ,chemistry.chemical_classification ,0303 health sciences ,Dipeptide ,Protease ,biology ,Lactococcus lactis ,biology.organism_classification ,Glucagon ,Fusion protein ,Recombinant Proteins ,Amino acid ,body regions ,Kinetics ,chemistry ,Biochemistry ,Amino Acid Substitution ,Proteolysis ,Mutagenesis, Site-Directed ,Biotechnology - Abstract
N-terminal extensions ("tags") have proven valuable for producing peptides using high throughput recombinant expression technologies. However, the applicability is hampered by the limited options for specific and efficient proteases to release the fully native sequence without additional amino acids in the N-terminal. Here we describe the Escherichia coli (E. coli) expression, purification and characterization of engineered variants of Xaa-Pro dipeptidyl aminopeptidase (Xaa-Pro-DAP) derived from Lactococcus lactis for cleavage of Gly-Pro dipeptide extension in the N-terminal of glucagon and glucagon-like peptide 1 (GLP-1(7-37)). By single amino acid substitution in the Xaa-Pro-DAP protease, significantly higher product yields were achieved. The combination of HRV14 3C protease and engineered Xaa-Pro-DAP is suggested for obtaining native N-terminal of peptides.
- Published
- 2019
48. Evaluation of serum prolidase activity and oxidative stress in patients with temporomandibular joint internal derangement
- Author
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Ismail Koyuncu, Mahmut Koparal, Ayşe Özcan Küçük, Bilal Ege, and Ataman Gönel
- Subjects
medicine.medical_specialty ,Dipeptidases ,Antioxidant ,medicine.medical_treatment ,Inflammation ,Temporomandibular joint internal derangement ,medicine.disease_cause ,Antioxidants ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,stomatognathic system ,Internal medicine ,medicine ,Humans ,In patient ,General Dentistry ,Temporomandibular Joint ,business.industry ,Significant difference ,030206 dentistry ,Glutathione ,Oxidative Stress ,Endocrinology ,Otorhinolaryngology ,chemistry ,Advanced oxidation protein products ,medicine.symptom ,business ,030217 neurology & neurosurgery ,Oxidative stress - Abstract
Objective: To investigate serum prolidase activity and oxidative stress in patients with temporomandibular joint internal derangement (TMJ-ID).Methods: Seventy patients with Wilkes stage III, IV, and V joints and 70 healthy controls were included. Serum prolidase activity and oxidative stress parameters, including total antioxidant status (TAS), total oxidant status (TOS), oxidative stress index (OSI), advanced oxidation protein products (AOPP), glutathione (GSH), ferric reducing antioxidant power (FRAP), and lipid hydroperoxide (LOOH) were measured.Results: The levels of prolidase, TOS, OSI, AOPP, and LOOH were significantly higher in the TMJ-ID group than in the control (p = .0001). TAS and FRAP level was significantly lower in the TMJ-ID group than in the control (p = .0001). There was no significant difference in GSH between groups.Conclusion: Significantly increased prolidase activity and oxidative stress in patients with TMJ-ID may be related to long-term collagen tissue damage, and inflammation and can be effective in the etiology of TMJ-ID.
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- 2019
49. A Novel Role of Prolidase in Cocaine-Mediated Breach in the Barrier of Brain Microvascular Endothelial Cells
- Author
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Jui Pandhare, Ife Albert, Fernando Villalta, Binah baht Ysrayl, Chandravanu Dash, and Muthukumar Balasubramaniam
- Subjects
0301 basic medicine ,Dipeptidases ,THP-1 Cells ,lcsh:Medicine ,Nitric Oxide Synthase Type II ,Pharmacology ,Blood–brain barrier ,Gene Expression Regulation, Enzymologic ,Monocytes ,Article ,03 medical and health sciences ,0302 clinical medicine ,Cocaine ,medicine ,Humans ,THP1 cell line ,lcsh:Science ,Regulation of gene expression ,Multidisciplinary ,biology ,Chemistry ,Monocyte ,lcsh:R ,Transendothelial and Transepithelial Migration ,Endothelial Cells ,Human brain ,3. Good health ,Endothelial stem cell ,Nitric oxide synthase ,030104 developmental biology ,medicine.anatomical_structure ,Blood-Brain Barrier ,Microvessels ,biology.protein ,Phosphorylation ,lcsh:Q ,030217 neurology & neurosurgery - Abstract
Cocaine use is associated with breach in the blood brain barrier (BBB) and increased HIV-1 neuro-invasion. We show that the cellular enzyme “Prolidase” plays a key role in cocaine-induced disruption of the BBB. We established a barrier model to mimic the BBB by culturing human brain microvascular endothelial cells (HBMECs) in transwell inserts. In this model, cocaine treatment enhanced permeability of FITC-dextran suggesting a breach in the barrier. Interestingly, cocaine treatment increased the activity of matrix metallo-proteinases that initiate degradation of the BBB-associated collagen. Cocaine exposure also induced prolidase expression and activity in HBMECs. Prolidase catalyzes the final and rate-limiting step of collagen degradation during BBB remodeling. Knock-down of prolidase abrogated cocaine-mediated increased permeability suggesting a direct role of prolidase in BBB breach. To decipher the mechanism by which cocaine regulates prolidase, we probed the inducible nitric oxide synthase (iNOS) mediated phosphorylation of prolidase since mRNA levels of the protein were not altered upon cocaine treatment. We observed increased iNOS expression concurrent with increased prolidase phosphorylation in cocaine treated cells. Subsequently, inhibition of iNOS decreased prolidase phosphorylation and reduced cocaine-mediated permeability. Finally, cocaine treatment increased transmigration of monocytic cells through the HBMEC barrier. Knock-down of prolidase reduced cocaine-mediated monocyte transmigration, establishing a key role of prolidase in cocaine-induced breach in endothelial cell barrier.
- Published
- 2019
50. Carnosine and Kidney Diseases: What We Currently Know?
- Author
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Katarzyna Kilis-Pstrusinska
- Subjects
Dipeptidases ,Carnosine ,Pharmacology ,Kidney ,Biochemistry ,Nephrotoxicity ,Proinflammatory cytokine ,Diabetic nephropathy ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Glycation ,Drug Discovery ,medicine ,Humans ,Diabetic Nephropathies ,030304 developmental biology ,0303 health sciences ,biology ,business.industry ,Organic Chemistry ,Acute Kidney Injury ,medicine.disease ,medicine.anatomical_structure ,chemistry ,Enzyme inhibitor ,030220 oncology & carcinogenesis ,biology.protein ,Molecular Medicine ,business ,Kidney disease - Abstract
Carnosine (beta-alanyl-L-histidine) is an endogenously synthesised dipeptide which is present in different human tissues e.g. in the kidney. Carnosine is degraded by enzyme serum carnosinase, encoding by CNDP1 gene. Carnosine is engaged in different metabolic pathways in the kidney. It reduces the level of proinflammatory and profibrotic cytokines, inhibits advanced glycation end products’ formation, moreover, it also decreases the mesangial cell proliferation. Carnosine may also serve as a scavenger of peroxyl and hydroxyl radicals and a natural angiotensin-converting enzyme inhibitor. : This review summarizes the results of experimental and human studies concerning the role of carnosine in kidney diseases, particularly in chronic kidney disease, ischemia/reperfusion-induced acute renal failure, diabetic nephropathy and also drug-induced nephrotoxicity. The interplay between serum carnosine concentration and serum carnosinase activity and polymorphism in the CNDP1 gene is discussed. : Carnosine has renoprotective properties. It has a promising potential for the treatment and prevention of different kidney diseases, particularly chronic kidney disease which is a global public health issue. Further studies of the role of carnosine in the kidney may offer innovative and effective strategies for the management of kidney diseases.
- Published
- 2019
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