19 results on '"Chang, Jeong Ho"'
Search Results
2. Morphology control of hierarchically ordered ceramic materials prepared by surfactant-directed sol-gel mineralization of wood cellular structures
- Author
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Shin, Yongsoon, primary, Wang, Li-Qiong, additional, Chang, Jeong Ho, additional, Samuels, William D., additional, and Exarhos, Gregory J., additional
- Published
- 2003
- Full Text
- View/download PDF
3. Targeting of CYP2E1 by miRNAs in alcohol-induced intestine injury.
- Author
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Mun H, Lee S, Choi S, Jeong JH, Ko S, Chun YL, Deaton B, Yeager CT, Boyette A, Palmera J, Newman L, Zhou P, Shin S, Kim DC, Sagum CA, Bedford MT, Kim YK, Kwon J, Jung J, Chang JH, and Yoon JH
- Subjects
- Humans, Animals, Heterogeneous Nuclear Ribonucleoprotein D0 metabolism, Intestines pathology, Intestinal Mucosa metabolism, Intestinal Mucosa pathology, Cytochrome P-450 CYP2E1 metabolism, Cytochrome P-450 CYP2E1 genetics, MicroRNAs metabolism, MicroRNAs genetics, Ethanol toxicity, Ethanol adverse effects
- Abstract
Although binge alcohol-induced gut leakage has been studied extensively in the context of reactive oxygen species-mediated signaling, it was recently revealed that post-transcriptional regulation plays an essential role as well. Ethanol (EtOH)-inducible cytochrome P450-2E1 (CYP2E1), a key enzyme in EtOH metabolism, promotes alcohol-induced hepatic steatosis and inflammatory liver disease, at least in part by mediating changes in intestinal permeability. For instance, gut leakage and elevated intestinal permeability to endotoxins have been shown to be regulated by enhancing CYP2E1 mRNA and CYP2E1 protein levels. Although it is understood that EtOH promotes CYP2E1 induction and activation, the mechanisms that regulate CYP2E1 expression in the context of intestinal damage remain poorly defined. Specific miRNAs, including miR-132, miR-212, miR-378, and miR-552, have been shown to repress the expression of CYP2E1, suggesting that these miRNAs contribute to EtOH-induced intestinal injury. Here, we have shown that CYP2E1 expression is regulated post-transcriptionally through miRNA-mediated degradation, as follows: (1) the RNA-binding protein AU-binding factor 1 (AUF1) binds mature miRNAs, including CYP2E1-targeting miRNAs, and this binding modulates the degradation of corresponding target mRNAs upon EtOH treatment; (2) the serine/threonine kinase mammalian Ste20-like kinase 1 (MST1) mediates oxidative stress-induced phosphorylation of AUF1. Those findings suggest that reactive oxygen species-mediated signaling modulates AUF1/miRNA interaction through MST1-mediated phosphorylation. Thus, our study demonstrates the critical functions of AUF1 phosphorylation by MST1 in the decay of miRNAs targeting CYP2E1, the stabilization of CYP2E1 mRNA in the presence of EtOH, and the relationship of this pathway to subsequent intestinal injury., Competing Interests: Declaration of Competing Interests The authors declare no competing interests., (Copyright © 2024. Published by Elsevier Inc.)
- Published
- 2024
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4. Structural basis for substrate recognition of glucose-6-phosphate dehydrogenase from Kluyveromyces lactis.
- Author
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Vu HH, Jin C, and Chang JH
- Subjects
- Amino Acid Sequence, Apoenzymes chemistry, Apoenzymes genetics, Apoenzymes metabolism, Binding Sites, Catalytic Domain, Crystallography, X-Ray, Glucosephosphate Dehydrogenase genetics, Kinetics, Models, Molecular, Mutagenesis, Structure-Activity Relationship, Substrate Specificity, Glucosephosphate Dehydrogenase chemistry, Glucosephosphate Dehydrogenase metabolism, Kluyveromyces enzymology
- Abstract
Glucose-6-phosphate dehydrogenase is the first enzyme in the pentose phosphate pathway. The reaction catalyzed by the enzyme is considered to be the main source of reducing power for nicotinamide adenine dinucleotide phosphate (NADPH) and is a precursor of 5-carbon sugar used by cells. To uncover the structural features of the enzyme, we determined the crystal structures of glucose-6-phosphate dehydrogenase from Kluyveromyces lactis (KlG6PD) in both the apo form and a binary complex with its substrate glucose-6-phosphate. KlG6PD contains a Rossman-like domain for cofactor NADPH binding; it also presents a typical antiparallel β sheet at the C-terminal domain with relatively the same pattern as those of other homologous structures. Moreover, our structural and biochemical analyses revealed that Lys153 contributes significantly to substrate G6P recognition. This study may provide insights into the structural variation and catalytic features of the G6PD enzyme., Competing Interests: Declaration of competing interest The authors have no conflict of interest to declare., (Copyright © 2021 Elsevier Inc. All rights reserved.)
- Published
- 2021
- Full Text
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5. Ursolic acid inhibits pigmentation by increasing melanosomal autophagy in B16F1 cells.
- Author
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Park HJ, Jo DS, Choi DS, Bae JE, Park NY, Kim JB, Chang JH, Shin JJ, and Cho DH
- Subjects
- Animals, Cell Line, Tumor, Melanins biosynthesis, Melanosomes drug effects, Mice, Triterpenes chemistry, alpha-MSH pharmacology, Ursolic Acid, Autophagy drug effects, Melanoma, Experimental pathology, Melanosomes pathology, Skin Pigmentation drug effects, Triterpenes pharmacology
- Abstract
Melanosomes are specialized membrane-bound organelles that are involved in melanin synthesis. Unlike melanosome biogenesis, the melanosome degradation pathway is poorly understood. Among the cellular processes, autophagy controls degradation of intracellular components by cooperating with lysosomes. In this study, we showed that ursolic acid inhibits skin pigmentation by promoting melanosomal autophagy, or melanophagy, in melanocytes. We found that B16F1 cells treated with ursolic acid suppressed alpha-melanocyte stimulating hormone (α-MSH) stimulated increase in melanin content and activated autophagy. In addition, we found that treatment with ursolic acid promotes melanosomal degradation, and bafilomycin A1 inhibition of autophagosome-lysosome fusion blocked the removal of melanosomes in α-MSH-stimulated B16F1 cells. Furthermore, depletion of the autophagy-related gene 5 (ATG5) resulted in significant suppression of ursolic acid-mediated anti-pigmentation activity and autophagy in α-MSH-treated B16F1 cells. Taken together, our results suggest that ursolic acid inhibits skin pigmentation by increasing melanosomal degradation in melanocytes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)
- Published
- 2020
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6. Corrigendum to "Transcriptional regulation of salmochelin glucosyltransferase by Fur in Salmonella".
- Author
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Lim D, Kim K, Song M, Jeong JH, Chang JH, Kim SR, Hong CW, Im SS, Park SH, Lee JC, and Shin M
- Published
- 2020
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7. Transcriptional regulation of Salmochelin glucosyltransferase by Fur in Salmonella.
- Author
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Lim D, Kim K, Song M, Jeong JH, Chang JH, Kim SR, Hong CW, Im SS, Park SH, Lee JC, and Shin M
- Subjects
- Bacterial Proteins chemistry, Base Sequence, Biobehavioral Sciences, Consensus Sequence, DNA, Bacterial genetics, Enterobactin metabolism, Gene Expression Regulation, Bacterial, Humans, Iron metabolism, Promoter Regions, Genetic, Protein Binding, Protein Multimerization, Repressor Proteins chemistry, Siderophores metabolism, Transcription, Genetic, Bacterial Proteins genetics, Bacterial Proteins metabolism, Enterobactin analogs & derivatives, Glucosyltransferases genetics, Glucosyltransferases metabolism, Repressor Proteins genetics, Repressor Proteins metabolism, Salmonella typhimurium genetics, Salmonella typhimurium metabolism
- Abstract
Pathogenic bacteria acquire the acquisition of iron from the host to ensure their survival. Salmonella spp. utilizes siderophores, including salmochelin, for high affinity aggressive import of iron. Although the iroBCDEN operon is reportedly responsible for the production and the transport of salmochelin, the molecular mechanisms underlying the regulation of its gene expression have not yet been characterized. Here, we analyzed the expression pattern of iroB using the lacZY transcriptional reporter system and determined the transcription start site in response to iron availability using primer extension analysis. We further examined the regulation of iroB expression by the ferric uptake regulator (Fur), a key regulatory protein involved in the maintenance of iron homeostasis in various bacteria, including Salmonella. Using sequence analysis followed by a gel shift assay, we verified that the Fur box lies within the promoter region of iroBCDE. The Fur box contained the consensus sequence (GATATTGGTAATTATTATC) and overlapped with the -10-element region. The expression of iroB was repressed by Fur in the presence of iron, as determined using an in vitro transcription assay. Therefore, we found that the iron acquisition system is regulated in a Fur-dependent manner in Salmonella., Competing Interests: Declaration of competing interest Authors have no conflicts of interest to declare., (Copyright © 2020 Elsevier Inc. All rights reserved.)
- Published
- 2020
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8. Loss of RNA binding protein, human antigen R enhances mitochondrial elongation by regulating Drp1 expression in SH-SY5Y cells.
- Author
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Bae JE, Park SJ, Hong Y, Jo DS, Lee H, Park NY, Kim JB, Park HJ, Bunch H, Chang JH, Lee EK, and Cho DH
- Subjects
- 1-Methyl-4-phenylpyridinium pharmacology, 3' Untranslated Regions genetics, Cell Line, Tumor, Dynamins metabolism, ELAV-Like Protein 1 metabolism, Herbicides pharmacology, Humans, Mitochondria drug effects, Mitochondria metabolism, Mitochondrial Dynamics drug effects, Protein Binding, RNA Interference, RNA, Messenger genetics, RNA, Messenger metabolism, RNA-Binding Proteins metabolism, Dynamins genetics, ELAV-Like Protein 1 genetics, Gene Expression Regulation, Neoplastic, Mitochondria genetics, Mitochondrial Dynamics genetics, RNA-Binding Proteins genetics
- Abstract
Mitochondria are essential for providing the energy necessary for neuronal function. Dysregulation of mitochondrial dynamics has been linked with the pathogenesis of many neurodegenerative diseases. Dynamin related protein 1 (Drp1) participates in fission activity in the mitochondria, and post-translational modifications to Drp1 modulate complex mitochondrial dynamics. However, the regulation of Drp1 at the post-transcriptional level remains poorly understood. In this study, we found that the RNA-binding protein Hu antigen R (HuR) post-transcriptionally regulates Drp1 expression. HuR interacts with Drp1 mRNA at its 3' untranslated region. Depletion of HuR reduces Drp1 expression, which leads to mitochondrial elongation in SH-SY5Y neuroblastoma cells. In contrast, ectopic expression of HuR enhances Drp1 expression, which promotes mitochondrial fragmentation in response to treatment with the mitochondrial complex 1 inhibitor MPP
+ . In addition, depletion of HuR suppressed the generation of mitochondrial ROS and cytotoxicity in MPP+ treated cells. Taken together, these findings suggest that HuR controls mitochondrial morphology via regulation of Drp1., (Copyright © 2019 Elsevier Inc. All rights reserved.)- Published
- 2019
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9. Crystal Structure of Histidine Triad Nucleotide-Binding Protein from the Pathogenic Fungus Candida albicans .
- Author
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Jung A, Yun JS, Kim S, Kim SR, Shin M, Cho DH, Choi KS, and Chang JH
- Subjects
- Amino Acid Sequence, Catalytic Domain, Crystallography, X-Ray, Humans, Phylogeny, Structural Homology, Protein, Candida albicans metabolism, Fungal Proteins chemistry, Hydrolases chemistry
- Abstract
Histidine triad nucleotide-binding protein (HINT) is a member of the histidine triad (HIT) superfamily, which has hydrolase activity owing to a histidine triad motif. The HIT superfamily can be divided to five classes with functions in galactose metabolism, DNA repair, and tumor suppression. HINTs are highly conserved from archaea to humans and function as tumor suppressors, translation regulators, and neuropathy inhibitors. Although the structures of HINT proteins from various species have been reported, limited structural information is available for fungal species. Here, to elucidate the structural features and functional diversity of HINTs, we determined the crystal structure of HINT from the pathogenic fungus Candida albicans (CaHINT) in complex with zinc ions at a resolution of 2.5 Å. Based on structural comparisons, the monomer of CaHINT overlaid best with HINT protein from the protozoal species Leishmania major . Additionally, structural comparisons with human HINT revealed an additional helix at the C-terminus of CaHINT. Interestingly, the extended C-terminal helix interacted with the N-terminal loop (α1-β1) and with the α3 helix, which appeared to stabilize the dimerization of CaHINT. In the C-terminal region, structural and sequence comparisons showed strong relationships among 19 diverse species from archea to humans, suggesting early separation in the course of evolution. Further studies are required to address the functional significance of variations in the C-terminal region. This structural analysis of CaHINT provided important insights into the molecular aspects of evolution within the HIT superfamily.
- Published
- 2019
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10. Effect of growth differentiation factor-15 secreted by human umbilical cord blood-derived mesenchymal stem cells on amyloid beta levels in in vitro and in vivo models of Alzheimer's disease.
- Author
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Kim DH, Lee D, Lim H, Choi SJ, Oh W, Yang YS, Chang JH, and Jeon HB
- Subjects
- Alzheimer Disease pathology, Animals, Coculture Techniques, Disease Models, Animal, Fetal Blood cytology, Growth Differentiation Factor 15 genetics, Growth Differentiation Factor 15 pharmacology, Humans, Insulysin metabolism, Mesenchymal Stem Cells cytology, Mice, Mutant Strains, Microglia drug effects, Microglia metabolism, Microglia pathology, Paracrine Communication, Peptide Fragments metabolism, Receptor, Transforming Growth Factor-beta Type II genetics, Receptor, Transforming Growth Factor-beta Type II metabolism, Recombinant Proteins genetics, Recombinant Proteins pharmacology, Alzheimer Disease metabolism, Amyloid beta-Peptides metabolism, Growth Differentiation Factor 15 metabolism, Mesenchymal Stem Cells metabolism
- Abstract
Alzheimer's disease (AD), which is the most common progressive neurodegenerative disease, causes learning and memory impairment. The pathological progress of AD can derive from imbalanced homeostasis of amyloid beta (Aβ) in the brain. In such cases, microglia play important roles in regulating the brain Aβ levels. In the present study, we found that human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) can increase, through paracrine action, the ability of microglial cells to clear Aβ. In order to identify the associated paracrine factors, a secretome of hUCB-MSCs co-cultured with Aβ-treated BV2 microglial cells was analyzed using a human cytokine protein array. As a result, growth differentiation factor-15 (GDF-15) was identified as a predominant candidate, and its association with Aβ clearance by microglial cells was investigated in vitro and in a 5XFAD mouse model. When Aβ-treated BV2 cells were treated with exogenous recombinant GDF-15, the Aβ levels in the culture medium decreased. Moreover, GDF-15 injection in the brain parenchyma of 5XFAD mice also led to decrease in Aβ plaques. In contrast, co-culture of BV2 cells and hUCB-MSCs treated with GDF-15-specific siRNA did not influence the Aβ levels in the culture medium. To elucidate how these phenomena are related, we confirmed that GDF-15 specifically increases insulin-degrading enzyme (IDE) expression in microglial cells through TGFβ receptor type II (TGFβRII), both in vitro and in vivo. These findings suggest that hUCB-MSCs promote the Aβ clearance ability of microglial cells through regulation of GDF-15 secretion, thus elucidating a therapeutic mechanism for AD., (Copyright © 2018 Elsevier Inc. All rights reserved.)
- Published
- 2018
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11. Molecular mechanism for the inhibition of DXO by adenosine 3',5'-bisphosphate.
- Author
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Yun JS, Yoon JH, Choi YJ, Son YJ, Kim S, Tong L, and Chang JH
- Subjects
- Adenosine Diphosphate metabolism, Binding Sites, Endoribonucleases antagonists & inhibitors, Endoribonucleases metabolism, Exoribonucleases metabolism, Magnesium chemistry, Models, Molecular, Adenosine Diphosphate chemistry, Endoribonucleases chemistry, Exoribonucleases chemistry
- Abstract
The decapping exoribonuclease DXO functions in pre-mRNA capping quality control, and shows multiple biochemical activities such as decapping, deNADding, pyrophosphohydrolase, and 5'-3' exoribonuclease activities. Previous studies revealed the molecular mechanisms of DXO based on the structures in complexes with a product, substrate mimic, cap analogue, and 3'-NADP
+ . Despite several reports on the substrate-specific reaction mechanism, the inhibitory mechanism of DXO remains elusive. Here, we demonstrate that adenosine 3', 5'-bisphosphate (pAp), a known inhibitor of the 5'-3' exoribonuclease Xrn1, inhibits the nuclease activity of DXO based on the results of structural and biochemical experiments. We determined the crystal structure of the DXO-pAp-Mg2+ complex at 1.8 Å resolution. In comparison with the DXO-RNA product complex, the position of pAp is well superimposed with the first nucleotide of the product RNA in the vicinity of two magnesium ions. Furthermore, biochemical assays showed that the inhibition by pAp is comparable between Xrn1 and DXO. Collectively, these structural and biochemical studies reveal that pAp inhibits the activities of DXO by occupying the active site to act as a competitive inhibitor., (Copyright © 2018 Elsevier Inc. All rights reserved.)- Published
- 2018
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- View/download PDF
12. eIF4E phosphorylation by MST1 reduces translation of a subset of mRNAs, but increases lncRNA translation.
- Author
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Min KW, Davila S, Zealy RW, Lloyd LT, Lee IY, Lee R, Roh KH, Jung A, Jemielity J, Choi EJ, Chang JH, and Yoon JH
- Subjects
- Amino Acid Sequence, Animals, Cell Line, Cell Line, Tumor, Gene Expression Regulation physiology, HeLa Cells, Humans, Mice, Polyribosomes metabolism, Protein Processing, Post-Translational physiology, RNA Caps metabolism, Signal Transduction physiology, Tumor Necrosis Factor-alpha metabolism, Eukaryotic Initiation Factor-4E metabolism, Hepatocyte Growth Factor metabolism, Phosphorylation physiology, Protein Biosynthesis physiology, Proto-Oncogene Proteins metabolism, RNA, Long Noncoding metabolism, RNA, Messenger metabolism
- Abstract
Post-transcriptional gene regulation is an important step in eukaryotic gene expression. The last step to govern production of nascent peptides is during the process of mRNA translation. mRNA translation is controlled by many translation initiation factors that are susceptible to post-translational modifications. Here we report that one of the translation initiation factors, eIF4E, is phosphorylated by Mammalian Ste20-like kinase (MST1). Upon phosphorylation, eIF4E weakly interacts with the 5' CAP to inhibit mRNA translation. Simultaneously, active polyribosome is more associated with long noncoding RNAs (lncRNAs). Moreover, the linc00689-derived micropeptide, STORM (Stress- and TNF-α-activated ORF Micropeptide), is triggered by TNF-α-induced and MST1-mediated eIF4E phosphorylation, which exhibits molecular mimicry of SRP19 and, thus, competes for 7SL RNA. Our findings have uncovered a novel function of MST1 in mRNA and lncRNA translation by direct phosphorylation of eIF4E. This novel signaling pathway will provide new platforms for regulation of mRNA translation via post-translational protein modification., (Published by Elsevier B.V.)
- Published
- 2017
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13. Crystal Structure and Comparative Sequence Analysis of GmhA from Colwellia psychrerythraea Strain 34H Provides Insight into Functional Similarity with DiaA.
- Author
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Do H, Yun JS, Lee CW, Choi YJ, Kim HY, Kim YJ, Park H, Chang JH, and Lee JH
- Subjects
- Bacterial Proteins chemistry, Bacterial Proteins genetics, Bacterial Proteins metabolism, Catalytic Domain, Cloning, Molecular, Crystallography, X-Ray, Gammaproteobacteria chemistry, Gammaproteobacteria genetics, Glutamine metabolism, Models, Molecular, Protein Conformation, Racemases and Epimerases metabolism, Structural Homology, Protein, Carrier Proteins chemistry, Gammaproteobacteria metabolism, Racemases and Epimerases chemistry, Racemases and Epimerases genetics, Sequence Analysis, Protein methods
- Abstract
The psychrophilic organism Colwellia psychrerythraea strain 34H produces extracellular polysaccharide substances to tolerate cold environments. Sedoheptulose 7-phosphate isomerase (GmhA) is essential for producing d-glycero-d-mannoheptose 7-phosphate, a key mediator in the lipopolysaccharide biosynthetic pathway. We determined the crystal structure of GmhA from C. psychrerythraea strain 34H (CpsGmhA, UniProtKB code: Q47VU0) at a resolution of 2.8 Å. The tetrameric structure is similar to that of homologous GmhA structures. Interestingly, one of the catalytic residues, glutamate, which has been reported to be critical for the activity of other homologous GmhA enzymes, is replaced by a glutamine residue in the CpsGmhA protein. We also found differences in the conformations of several other catalytic residues. Extensive structural and sequence analyses reveal that CpsGmhA shows high similarity to Escherichia coli DnaA initiator-associating protein A (DiaA). Therefore, the CpsGmhA structure reported here may provide insight into the structural and functional correlations between GmhA and DiaA among specific microorganisms.
- Published
- 2015
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14. Structural basis for the ligand-binding specificity of fatty acid-binding proteins (pFABP4 and pFABP5) in gentoo penguin.
- Author
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Lee CW, Kim JE, Do H, Kim RO, Lee SG, Park HH, Chang JH, Yim JH, Park H, Kim IC, and Lee JH
- Subjects
- Amino Acid Sequence, Anilino Naphthalenesulfonates, Animals, Avian Proteins genetics, Avian Proteins metabolism, Cloning, Molecular, Crystallography, X-Ray, Escherichia coli genetics, Escherichia coli metabolism, Fatty Acid-Binding Proteins genetics, Fatty Acid-Binding Proteins metabolism, Fluorescent Dyes, Gene Expression, Ligands, Models, Molecular, Molecular Sequence Data, Protein Binding, Protein Isoforms chemistry, Protein Isoforms genetics, Protein Isoforms metabolism, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Sequence Alignment, Sequence Homology, Amino Acid, Spheniscidae genetics, Avian Proteins chemistry, Fatty Acid-Binding Proteins chemistry, Linoleic Acid chemistry, Palmitic Acid chemistry, Spheniscidae metabolism
- Abstract
Fatty acid-binding proteins (FABPs) are involved in transporting hydrophobic fatty acids between various aqueous compartments of the cell by directly binding ligands inside their β-barrel cavities. Here, we report the crystal structures of ligand-unbound pFABP4, linoleate-bound pFABP4, and palmitate-bound pFABP5, obtained from gentoo penguin (Pygoscelis papua), at a resolution of 2.1 Å, 2.2 Å, and 2.3 Å, respectively. The pFABP4 and pFABP5 proteins have a canonical β-barrel structure with two short α-helices that form a cap region and fatty acid ligand binding sites in the hydrophobic cavity within the β-barrel structure. Linoleate-bound pFABP4 and palmitate-bound pFABP5 possess different ligand-binding modes and a unique ligand-binding pocket due to several sequence dissimilarities (A76/L78, T30/M32, underlining indicates pFABP4 residues) between the two proteins. Structural comparison revealed significantly different conformational changes in the β3-β4 loop region (residues 57-62) as well as the flipped Phe60 residue of pFABP5 than that in pFABP4 (the corresponding residue is Phe58). A ligand-binding study using fluorophore displacement assays shows that pFABP4 has a relatively strong affinity for linoleate as compared to pFABP5. In contrast, pFABP5 exhibits higher affinity for palmitate than that for pFABP4. In conclusion, our high-resolution structures and ligand-binding studies provide useful insights into the ligand-binding preferences of pFABPs based on key protein-ligand interactions., (Copyright © 2015 Elsevier Inc. All rights reserved.)
- Published
- 2015
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15. Crystal structure and biochemical properties of the (S)-3-hydroxybutyryl-CoA dehydrogenase PaaH1 from Ralstonia eutropha.
- Author
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Kim J, Chang JH, and Kim KJ
- Subjects
- 3-Hydroxyacyl CoA Dehydrogenases genetics, Acyl Coenzyme A chemistry, Acyl Coenzyme A metabolism, Amino Acid Sequence, Crystallography, X-Ray, Kinetics, Models, Molecular, Molecular Sequence Data, Mutagenesis, Site-Directed, NAD chemistry, NAD metabolism, Protein Conformation, 3-Hydroxyacyl CoA Dehydrogenases chemistry, 3-Hydroxyacyl CoA Dehydrogenases metabolism, Cupriavidus necator enzymology
- Abstract
3-Hydroxybutyryl-CoA dehydrogenase is an enzyme involved in the synthesis of the biofuel n-butanol by converting acetoacetyl-CoA to 3-hydroxybutyryl-CoA. To investigate the molecular mechanism of n-butanol biosynthesis, we determined crystal structures of the Ralstonia eutropha-derived 3-hydroxybutyryl-CoA dehydrogenase (RePaaH1) in complex with either its cofactor NAD(+) or its substrate acetoacetyl-CoA. While the biologically active structure is dimeric, the monomer of RePaaH1 comprises two separated domains with an N-terminal Rossmann fold and a C-terminal helical bundle for dimerization. In this study, we show that the cofactor-binding site is located on the Rossmann fold and is surrounded by five loops and one helix. The binding mode of the acetoacetyl-CoA substrate was found to be that the adenosine diphosphate moiety is not highly stabilized compared with the remainder of the molecule. Residues involved in catalysis and substrate binding were further confirmed by site-directed mutagenesis experiments, and kinetic properties of RePaaH1were examined as well. Our findings contribute to the understanding of 3-hydroxybutyryl-CoA dehydrogenase catalysis, and will be useful in enhancing the efficiency of n-butanol biosynthesis by structure based protein engineering., (Copyright © 2014 Elsevier Inc. All rights reserved.)
- Published
- 2014
- Full Text
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16. Crystal structure of (R)-3-hydroxybutyryl-CoA dehydrogenase PhaB from Ralstonia eutropha.
- Author
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Kim J, Chang JH, Kim EJ, and Kim KJ
- Subjects
- 3-Hydroxyacyl CoA Dehydrogenases metabolism, Amino Acid Sequence, Coenzymes metabolism, Crystallography, X-Ray, Enzyme Assays, Models, Molecular, Molecular Sequence Data, Mutagenesis, Mutant Proteins chemistry, Mutant Proteins metabolism, NADP metabolism, Protein Structure, Tertiary, Substrate Specificity, 3-Hydroxyacyl CoA Dehydrogenases chemistry, Cupriavidus necator enzymology
- Abstract
(R)-3-hydroxybutyryl-CoA dehydrogenase PhaB from Ralstonia eutropha H16 (RePhaB) is an enzyme that catalyzes the NADPH-dependent reduction of acetoacetyl-CoA, an intermediate of polyhydroxyalkanoates (PHA) synthetic pathways. Polymeric PHA is used to make bioplastics, implant biomaterials, and biofuels. Here, we report the crystal structures of RePhaB apoenzyme and in complex with either NADP(+) or acetoacetyl-CoA, which provide the catalytic mechanism of the protein. RePhaB contains a Rossmann fold and a Clamp domain for binding of NADP(+) and acetoacetyl-CoA, respectively. The NADP(+)-bound form of RePhaB structure reveals that the protein has a unique cofactor binding mode. Interestingly, in the RePhaB structure in complex with acetoacetyl-CoA, the conformation of the Clamp domain, especially the Clamp-lid, undergoes a large structural change about 4.6 Å leading to formation of the substrate pocket. These structural observations, along with the biochemical experiments, suggest that movement of the Clamp-lid enables the substrate binding and ensures the acetoacetyl moiety is located near to the nicotinamide ring of NADP(+)., (Copyright © 2013 Elsevier Inc. All rights reserved.)
- Published
- 2014
- Full Text
- View/download PDF
17. Amino acid side chain-like surface modification on magnetic nanoparticles for highly efficient separation of mixed proteins.
- Author
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Lee SY, Ahn CY, Lee J, and Chang JH
- Subjects
- Animals, Cattle, Silicon Dioxide chemistry, Solutions, Surface Properties, Amino Acids chemistry, Chemical Fractionation methods, Magnetite Nanoparticles chemistry, Muramidase chemistry, Muramidase isolation & purification, Serum Albumin, Bovine chemistry, Serum Albumin, Bovine isolation & purification
- Abstract
This work reports on the realization of specific functionalized silica-coated magnetic nanoparticles (Si-MNPs) for effective protein separation through surface modification with various amino acid side chain-like functional groups such as thiol (-SH), disulfide (-S-S-), carbon chain (-C(n)), carboxyl (-COOH), amine (-NH(2)), and aldehyde (-CHO). This study also suggests an improved and convenient method for the synthesis of functionalized Si-MNPs by hydrolysis condensation with silan-coupling agents. The protein adsorption effects in a coexistent mixed state are explored using various proteins, which have different isoelectric point (pI) values and molecular weights, in order to elucidate the binding performance of different proteins one solution. The adsorption efficiency of bovine serum albumin (BSA; 66 kDa; pI=4.65) and lysozyme (LYZ; 14.3 kDa; pI=11) is 70-100% with various amino acid side chain-like functional groups. However, the adsorption efficiency of a mixed protein solution of BSA and LYZ was different. Although the relatively bulky BSA molecule displayed 50% and 20% adsorption corresponding to pH 4.65, and pH 11, respectively, the smaller LYZ provided almost 100% adsorption at both pH 4.65 and pH 11., (Copyright © 2012 Elsevier B.V. All rights reserved.)
- Published
- 2012
- Full Text
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18. Optimization of biotin labeling of antibodies using mouse IgG and goat anti-mouse IgG-conjugated fluorescent beads and their application as capture probes on protein chip.
- Author
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Lee JH, Choi HK, and Chang JH
- Subjects
- Animals, Antibodies, Immobilized immunology, Avidin chemistry, Avidin immunology, Biotin immunology, Fluorescent Dyes, Goats, Humans, Immunoassay methods, Immunoglobulin G immunology, Mice, Natriuretic Peptide, Brain immunology, Peptide Fragments immunology, Troponin I immunology, Antibodies, Immobilized chemistry, Biotin chemistry, Immunoglobulin G chemistry, Natriuretic Peptide, Brain analysis, Peptide Fragments analysis, Protein Array Analysis methods, Troponin I analysis
- Abstract
This study shows the optimization of biotin labeling to antibodies using mouse IgG. Several parameters of the biotin labeling, including the molar ratio of biotin to antibody, the coupling time and the dialysis time, were studied to optimum conditions. The biotin-tagged mouse IgGs were immobilized on avidin-coated PMMA (Polymethyl Methacrylate) plates via a biotin-avidin linkage. The immobilization of the IgG to the chip was quantified using goat anti-mouse IgG bound fluorescent beads. It was found that the binding of the fluorescent beads saturated when a 10-fold or higher molar ratio of biotin to antibody was used. In biotin coupling time tests, sixty minutes was sufficient for the capture probes to bind to the surface. However, the results from the dialysis experiments showed no difference, indicating that 2 hours was sufficient to remove any unbound biotin. Finally, to prove the universality of this protocol using mouse antibodies, the optimum conditions were successfully applied in sandwich immunoassays designed to detect troponin I (TnI) and N-terminal probrain natriuretic peptide (NT-proBNP)., (Crown Copyright © 2010. Published by Elsevier B.V. All rights reserved.)
- Published
- 2010
- Full Text
- View/download PDF
19. Hydrophobic partitioning approach to efficient protein separation with magnetic nanoparticles.
- Author
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Chang JH, Lee J, Jeong Y, Hyung Lee J, Kim IJ, and Park SE
- Subjects
- Adsorption, Animals, Cattle, Hydrogen-Ion Concentration, Hydrophobic and Hydrophilic Interactions, Silicon Dioxide chemistry, Chromatography, Liquid methods, Magnetics, Nanoparticles chemistry, Serum Albumin, Bovine chemistry
- Abstract
A novel approach to the hydrophobic partitioning effect on efficient separation of protein such as BSA was demonstrated by the modification of hydrophobic pockets on the surface of silica-coated magnetic nanoparticles with various alkyl groups at various pH levels. The separation efficiency is strongly reflected and can be attained by controlling the size of the hydrophobic pocket and other factors such as the alkyl chain length, the salt concentration, and the pH levels. Furthermore, the adsorption constant (k(ad)) was calculated for the hydrophobic partitioning interaction between BSA and the hydrophobic pocket size at various pH levels., (Copyright 2010 Elsevier Inc. All rights reserved.)
- Published
- 2010
- Full Text
- View/download PDF
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