Search

In this text, we discuss the filter banks used for speech analysis and propose a novel filter bank for speech processing applications. Filter banks are building blocks of speech processing applications. Multiple filter strategies have been proposed, including Mel, PLP, Seneff, Lyon, and Gammatone filters. MFCC is a transformed version of Mel filters and is still a state-of-the-art method for speech recognition applications. However, 40 years after their debut, time is running out to launch new structures as novel speech features. The proposed acoustic filter banks (AFB) are innovative alternatives to dethrone Mel filters, PLP filters, and MFCC features. Foundations of AFB filters are based on the formant regions of vowels and consonants. In this study, we pioneer an acoustic filter bank comprising 11 frequency regions and conduct experiments using the VGG16 model on the TIMIT and Speech Command V2 datasets. The outcomes of the study concretely indicate that MFCC, Mel, and PLP filters can effectively be replaced with novel AFB filter bank features. [ABSTRACT FROM AUTHOR]

l-Homocysteine, formed from S-adenosyl methionine following demethylation and adenosine release, accumulates when the methionine recycling pathway and other pathways become impaired, thus leading to hyperhomocysteinemia, a biomarker in cardiovascular diseases, neurological/psychiatric disorders, and cancer. The partial oxidation of the l-homocysteine thiol group and its decarboxylation on C-alpha lead to the formation of l-homocysteinesulfinic acid (l-HCSA) and homohypotaurine (HHT), respectively. Both compounds are not readily available from commercial suppliers, which hinders the investigation of their biological activities. Herein, the chemical synthesis of l-HCSA, from l-homocystine, was the starting point for establishing the bio-based synthesis of HHT using recombinant Escherichia coli glutamate decarboxylase (EcGadB), an enzyme already successfully employed for the bio-based synthesis of GABA and its phosphinic analog. Prior to HHT synthesis, kcat (33.92 ± 1.07) and KM (38.24 ± 3.45 mM) kinetic constants were determined for l-HCSA on EcGadB. The results of our study show that the EcGadB-mediated synthesis of HHT can be achieved with good yields (i.e., 40% following enzymatic synthesis and column chromatography). Purified HHT was tested in vitro on primary human umbilical vein endothelial cells and rat cardiomyoblasts and compared to the fully oxidized analog, homotaurine (OT, also known as tramiprosate), in widespread pharmaceutical use. The results show that both cell lines display statistically significant recovery from the cytotoxic effects induced by H2O2 in the presence of HHT. [ABSTRACT FROM AUTHOR]

Aim: The aim of this study was to investigate the role of pyridoxal-5-phosphate (PLP) level on the oral health status as a predictive marker in patients with hypophosphatasia (HPP). Materials and methods: Throughout a systematic retrospective assessment both bone metabolism and oral health status were analyzed. The oral health status was assessed by the decayed/missing/filled teeth index (DMFT), clinical attachment level (CAL), probing pocket depth (PPD), and the periodontal screening index (PSI). Results: A total of 48 HPP patients (81.3% female) with a mean age of 42.21 years was included in this retrospective study. The study population was divided into two groups using the mean PLP level (87 µg/l) as a cut-off. Patients with a PLP level ≥ 87 µg/l (n = 14) showed a significantly poorer oral health status regarding DMFT index, CAL, PPD and PSI compared to patients with a PLP level < 87 µg/l (n = 34). No significant group differences for tooth loss were found. Conclusion: The results of the present study indicate that the PLP level is a suitable diagnostic predictor for the oral health status in HPP patients. HPP patients with PLP levels ≥ 70 µg/l should be included into a regular dental preventive program. Clinical Relevance: The oral health status in HPP and its correlation with laboratory parameters (i.e. PLP) has been understudied. For clinical practice, the findings of the present study clearly demonstrated that high PLP levels correlate with a worse oral health status in HPP patients. Therefore, these patients should receive an intensive dental treatment and/or inclusion in a strict maintenance program in a specialized dental practice/university hospital with a PLP level ≥ 70 µg/l. [ABSTRACT FROM AUTHOR]

1,4-diaminobutane is widely used in the industrial production of polymers, pharmaceuticals, agrochemicals and surfactants. Owing to economic and environmental concerns, there has been a growing interest in using microbes to produce 1,4-diaminobutane. However, there is lack of research on the influence of cofactors pyridoxal phosphate (PLP) and NADPH on the synthesis of 1,4-diaminobutane. PLP serves as a cofactor of ornithine decarboxylase in the synthesis of 1,4- diaminobutane. Additionally, the synthesis of 1 mol 1,4-diaminobutane requires 2 mol NADPH, thus necessitating consideration of NADPH balance in the efficient synthesis of 1,4-diaminobutane by Escherichia coli. The aim of this study was to enhance the synthesis efficiency of 1,4-diaminobutane through increasing production of PLP and NADPH. By optimizing the expression of the genes associated with synthesis of PLP and NADPH in E. coli, cellular PLP and NADPH levels increased, and the yield of 1,4-diaminobutane also increased accordingly. Ultimately, using glucose as the primary carbon source, the yield of 1,4-diaminobutane in the recombinant strain NAP19 reached 272 mg/L·DCW, by increased 79% compared with its chassis strain. [ABSTRACT FROM AUTHOR]

The development of medical technology devices leads to the introduction and use of agile methods, which enable the delivery of increasingly complex software with the fastest possible innovations. Delivery of the highest quality software must be considered during development, as medical products are important elements in saving human lives. Their development begins with determining a set of product requirements that exactly correspond to it. The development of specified medical products is finally delivered to the customer, who participates in the development. In this article, we present the use and combination of agile methods in software development, which correct and facilitate timely and continuous delivery of products. They also know how to smooth out a quick reaction to the customer's changing needs and mainly focus on team management and communication. Specific agile methods make it possible to implement development through gradual improvements by integrating customer requirements towards the product. This article identifies three interconnected approaches to integrating agile methods and principles: SCRUM, SAFe, and Kanban combined with the V-model. The methods are gradually analysed based on the literature review, and the article presents a practical application in Siemens Healthcare Slovakia. [ABSTRACT FROM AUTHOR]

Pyridoxal 5´-phosphate (PLP) is the main form of vitamin B6 in animal tissue and functions as a coenzyme for more than 160 different enzymatic reactions in the metabolism of amino acids, carbohydrates, lipids, and neurotransmitters. Estimated dietary intake of vitamin B6 and plasma PLP values differ a lot between studies, something which may be due to variable use of supplements, variations in dietary assessment and analytical methods. These factors make it difficult to achieve precise data for setting a correct recommended intake of vitamin B6. In addition, a plasma PLP concentration of 30 nmol/L is considered to be sufficient and the current recommendations for vitamin B6 intake is based on this concept. However, the metabolic marker for vitamin B6 status, HK ratio (HKr), starts to increase already when plasma PLP falls below 100 nmol/L and increases more steeply below 50 nmol/L, indicating biochemical deficiency. Consequently, a plasma PLP concentration of 30 nmol/L, may be too low as a marker for an adequate vitamin B6 status.

The serine peptidase CLPP is conserved among bacteria, chloroplasts, and mitochondria. In humans and mice, its loss causes Perrault syndrome, which presents with growth deficits, infertility, deafness, and ataxia. In the filamentous fungus Podospora anserina, CLPP loss leads to longevity. CLPP substrates are selected by CLPX, an AAA+ unfoldase. CLPX is known to target delta-aminolevulinic acid synthase (ALAS) to promote pyridoxal phosphate (PLP) binding. CLPX may also influence cofactor association with other enzymes. Here, the evaluation of P. anserina metabolomics highlighted a reduction in arginine/histidine levels. In Mus musculus cerebellum, reductions in arginine/histidine and citrulline occurred with a concomitant accumulation of the heme precursor protoporphyrin IX. This suggests that the increased biosynthesis of 5-carbon (C5) chain deltaALA consumes not only C4 succinyl-CoA and C1 glycine but also specific C5 delta amino acids. As enzymes responsible for these effects, the elevated abundance of CLPX and ALAS is paralleled by increased OAT (PLP-dependent, ornithine delta-aminotransferase) levels. Possibly as a consequence of altered C1 metabolism, the proteome profiles of P. anserina CLPP-null cells showed strong accumulation of a methyltransferase and two mitoribosomal large subunit factors. The reduced histidine levels may explain the previously observed metal interaction problems. As the main nitrogen-storing metabolite, a deficiency in arginine would affect the urea cycle and polyamine synthesis. Supplementation of arginine and histidine might rescue the growth deficits of CLPP-mutant patients. [ABSTRACT FROM AUTHOR]

Speech is one of the communication processes of humans. One of the important features of speech is to convey the inner feelings of the person to the listener. When a speech is expressed by the speaker, this speech also contains the feelings of the person, which leads to the creation of thoughts and behaviors appropriate to oneself. Speech Emotion Recognition (SER) is a very important issue in the field of human–machine interaction. The expansion of the use of computers and its impact on today's life has caused this mutual cooperation between man and machine to be widely investigated and researched. In this article, SER in English and Persian has been examined. Frequency time characteristics such as Mel- Frequency Cepstral Coefficient (MFCC), Linear Predictive Coding and Predictive Linear Perceptual (PLP) are extracted from the data as feature vectors, then they are combined with each other and a selection of suitable features from them. Also, Principal components analysis (PCA) is used to reduce dimensions and eliminate redundancy while retaining most of the intrinsic information content of the pattern. Then, each emotional state was classified using the Gaussian Mixtures Model (GMM) and Hidden Markov Model (HMM) technique. Combining the MFCC + PLP properties, PCA features, and HMM classification with a precision of 88.85% and a runtime of 0.3 s produces the average diagnostic rate in the English database; similarly, the PLP properties, PCA features, and HMM classification with a precision of 90.21% and a runtime of 0.4 s produce the average diagnostic rate in the Persian database. Based on the combination of features and classifications, the experimental results demonstrated that the suggested approach can attain a high level of stable detection performance for every emotional state. [ABSTRACT FROM AUTHOR]

The reproductive efficiency improvements that can be achieved in bovine herds highly depend on the male, as the bulls account for approximately 80% of the gene pool. Likewise, age can impact the sires' reproductive efficiency through its seminal quality, which is strongly influenced by the bull's sexual maturity. The study aimed to assess the bull's age effect on the X and Y chromosomes-bearing spermatozoa ratio in semen. Sixteen Brahman bulls (Bos indicus) were grouped into two age range and their reproductive capacity was assessed. Genomic DNA was extracted from a single collection of semen to quantify by qPCR (quantitative polymerase chain reaction) the PLP and SRY genes, used as sexual chromosome markers, and autosomal GAPDH as the reference gene. The values obtained were compared using the delta Ct (2-Ct) method and validated with the use of a commercial sample of semen sexed for the X chromosome. The SRY gene had a proportion of 43% in bulls of 4-5 years of age and 32% in the group of 9-10 years, while the proportions of the PLP gene were 57% in bulls of 4-5 years and 68% in the 9-10 years group. This deviation from the expected ratio (1:1) in bulls of nine and ten years old with a preponderance of the X chromosome content might suggest a possible influence of age in the sex chromosome proportions in the spermatozoa; however, these results were not statistically significant. [ABSTRACT FROM AUTHOR]

This paper introduces an innovative technique for creating a cough detection system that relies on speech recognition algorithms. The strategy utilizes the Kaldi platform, which is open source and incorporates a hybrid system of Gaussian Mixture Model-based Hidden Markov Models (GMM-HMM) through a straightforward monophone training model. Additionally, the study examines the effectiveness of two different feature extraction approaches, Mel Frequency Cepstral Coefficient (MFCC) and Perceptual Linear Prediction (PLP). The proposed system can function as a collection tool for gathering natural and spontaneous cough data from conversations or continuous speech. The paper also compares the Kaldi and CMU Sphinx4 toolkits, concluding that Kaldi's use of GMM-HMM outperforms CMU Sphinx4. [ABSTRACT FROM AUTHOR]

l-Homocysteine, formed from S-adenosyl methionine following demethylation and adenosine release, accumulates when the methionine recycling pathway and other pathways become impaired, thus leading to hyperhomocysteinemia, a biomarker in cardiovascular diseases, neurological/psychiatric disorders, and cancer. The partial oxidation of the l-homocysteine thiol group and its decarboxylation on C-alpha lead to the formation of l-homocysteinesulfinic acid (l-HCSA) and homohypotaurine (HHT), respectively. Both compounds are not readily available from commercial suppliers, which hinders the investigation of their biological activities. Herein, the chemical synthesis of l-HCSA, from l-homocystine, was the starting point for establishing the bio-based synthesis of HHT using recombinant Escherichia coli glutamate decarboxylase (EcGadB), an enzyme already successfully employed for the bio-based synthesis of GABA and its phosphinic analog. Prior to HHT synthesis, kcat (33.92 ± 1.07) and KM (38.24 ± 3.45 mM) kinetic constants were determined for l-HCSA on EcGadB. The results of our study show that the EcGadB-mediated synthesis of HHT can be achieved with good yields (i.e., 40% following enzymatic synthesis and column chromatography). Purified HHT was tested in vitro on primary human umbilical vein endothelial cells and rat cardiomyoblasts and compared to the fully oxidized analog, homotaurine (OT, also known as tramiprosate), in widespread pharmaceutical use. The results show that both cell lines display statistically significant recovery from the cytotoxic effects induced by H2O2 in the presence of HHT.

Based on the phenomenon that happened to Surabaya State University students who had implemented the Pengenalan Lapangan Persekolahan (PLP) program, it showed that there was a lack of interest in becoming a teacher. One of the internal factors is teaching experience in the form of the PLP program and the external factor is the family environment. Goal of the study was to analyze the impact of school district (PLP) introduction and home environment on the educational interests of students in administrative education at Surabaya State University. This kind of examination is quantitative using explanatory. The subjects of this research were all students of the Surabaya State University Office Administration Study Program class of 2019 and then had taken PLP courses with a total of 54 people. The instrument used was a questionnaire with a Likert scale of 5. Data analysis techniques include testing for validity also reliability. Classical acceptance tests consist of normality, multicollinearity, also heteroscedasticity tests, and include hypothesis testing using several linear regression tests consisting of t-tests, F-tests, and coefficient-of-squares tests. The outcome of the t-test showed that the introduction of school fields had a positive and significant effect on interest in becoming a teacher, and the family environment had a positive and significant effect on interest in becoming a teacher. The introduction of school fields and family environment has had a positive and significant impact on students’ savor in becoming a teacher in University Office Administration Education students Surabaya Country.

Effective learning is an important aspect for students as a provision for the introduction of field schooling (PLP) for students. The purpose of this study was to determine how much students can apply effective learning models in the learning process when following PLP, as well as how the application of these models affect the effectiveness of learning when following PLP. The design of research conducted in this study is qualitative research with descriptive approach. The population in this study as many as 55 students and then sampled a number of 30 students with random sampling. The results of the observation sheet showed that most students (70%) can apply effective learning models in the learning process when following PLP. While the other 30% still have difficulty in implementing the model. The results of data reduction from interview sheets showed that most students (80%) stated that the application of effective learning models in PLP makes learning more fun and effective. Another 20% stated that they had difficulties in implementing the model. The results of data reduction from the questionnaire showed that most students choose expository learning (20%) Problem-Based (60%) and cooperative learning (15%), most students can apply problem-based learning models in the learning process when attending PLP. This can be seen from the reduction of observation sheet data which shows that 70% of students can apply the model. The results of interviews and questionnaires also showed that most students stated that the application of effective learning models in PLP makes learning more fun and effective.

Penelitian ini bertujuan untuk mengetahui pengaruh Pengenalan Pendidikan Sekolah (PLP), KKN-Dik, dan penguasaan teknologi informasi terhadap minat menjadi guru akuntansi di era Society 5.0. Penelitian ini merupakan penelitian kuantitatif dengan menggunakan metode pengumpulan data kuesioner. Kuesioner disebarkan kepada 129 sampel dengan menggunakan teknik purposive sampling. Metode analisis data pada penelitian ini menggunakan analisis regresi linier berganda yang sebelumnya diuji asumsi klasik dengan menggunakan SPSS for Windows Versi 23. Berdasarkan hasil uji t diperoleh nilai signifikansi hasil analisis variabel PLP sebesar 0.001, KKN-Dik adalah 0.030, dan kemampuan teknologi informasi 0.000. Artinya ketiga variabel tersebut mempunyai nilai penting sehingga secara parsial mempengaruhi minat menjadi guru akuntansi. Sedangkan hasil uji F diperoleh nilai signifikan 0,000. Hal ini menunjukkan bahwa PLP, KKN-Dik, dan penguasaan teknologi informasi secara simultan berpengaruh terhadap minat menjadi guru akuntansi di era society 5.0. Disamping uji t dan uji F, nilai R square (R2 ) sebesar 0, 329 artinya 32,9% minat menjadi guru akuntansi dipengaruhi oleh pengenalan lapangan persekolahan (PLP), KKN-Dik, dan penguasaan teknologi infromasi. Sedangkan 67,1% dapat dijelaskan oleh faktor lain yang tidak diteliti dalam penelitian ini.

Reduction in dietary vitamin B6 intake is associated with an increased relative risk of diseases such as cancer, atherosclerosis and cognitive dysfunction. The current research has assessed vitamin B6 intakes and PLP concentrations as a marker of vitamin B6 status among the UK adult (≥ 19 years) population. This study was carried out using a cross-sectional analysis of the National Diet and Nutrition Survey Rolling Programme (NDNS) (2008–2017). The impacts of lifestyle factors, including type of diet, smoking, alcohol consumption, and commonly used medications grouped by therapeutic usage, were determined, and data were analysed using IBM SPSS®. Results are expressed as medians (25th–75th percentiles), with P values ≤ 0·05 considered statistically significant. Among UK adults, the median intakes of total population of dietary vitamin B6 met the reference nutrient intake and median plasma PLP concentrations were above the cut-off of vitamin B6 deficiency; however, we found an association between reduction in vitamin B6 intake and plasma PLP concentration and age group (P < 0·001). Smokers had significantly lower plasma PLP concentrations than non-smokers (P < 0·001). Moreover, regression analysis showed some commonly used medications were associated with plasma PLP levels reduction (P < 0·05). Taken together, we report on a tendency for dietary vitamin B6 intake and plasma PLP concentrations to decrease with age and lifestyle factors such as smoking and medication usage. This information could have important implications for smokers and in the elderly population using multiple medications (polypharmacy). [ABSTRACT FROM AUTHOR]

We previously demonstrated that people with primary sclerosing cholangitis (PSC) had reduced gut microbial capacity to produce active vitamin B6 (pyridoxal 5'-phosphate [PLP]), which corresponded to lower circulating PLP levels and poor outcomes. Here, we define the extent and biochemical and clinical impact of vitamin B6 deficiency in people with PSC from several centers before and after liver transplantation (LT). We used targeted liquid chromatography-tandem mass spectrometry to measure B6 vitamers and B6-related metabolic changes in blood from geographically distinct cross-sectional cohorts totaling 373 people with PSC and 100 healthy controls to expand on our earlier findings. Furthermore, we included a longitudinal PSC cohort (n = 158) sampled prior to and serially after LT, and cohorts of people with inflammatory bowel disease (IBD) without PSC (n = 51) or with primary biliary cholangitis (PBC) (n = 100), as disease controls. We used Cox regression to measure the added value of PLP to predict outcomes before and after LT. In different cohorts, 17-38% of people with PSC had PLP levels below the biochemical definition of a vitamin B6 deficiency. The deficiency was more pronounced in PSC than in IBD without PSC and PBC. Reduced PLP was associated with dysregulation of PLP-dependent pathways. The low B6 status largely persisted after LT. Low PLP independently predicted reduced LT-free survival in both non-transplanted people with PSC and in transplant recipients with recurrent disease. Low vitamin B6 status with associated metabolic dysregulation is a persistent feature of PSC. PLP was a strong prognostic biomarker for LT-free survival both in PSC and recurrent disease. Our findings suggest that vitamin B6 deficiency modifies the disease and provides a rationale for assessing B6 status and testing supplementation. We previously found that people with PSC had reduced gut microbial potential to produce essential nutrients. Across several cohorts, we find that the majority of people with PSC are either vitamin B6 deficient or have a marginal deficiency, which remains prevalent even after liver transplantation. Low vitamin B6 levels strongly associate with reduced liver transplantation-free survival as well as deficits in biochemical pathways dependent on vitamin B6, suggesting that the deficiency has a clinical impact on the disease. The results provide a rationale for measuring vitamin B6 and to investigate whether vitamin B6 supplementation or modification of the gut microbial community can help improve outcomes for people with PSC. [Display omitted] • Vitamin B6 deficiency is common in people with primary sclerosing cholangitis. • Low vitamin B6 impaired PLP-dependent pathways important for health. • Low vitamin B6 was associated with short liver transplantation-free survival. • The addition of vitamin B6 to current risk models improved survival prediction. • After liver transplantation, low B6 persisted and was associated with poor outcomes. [ABSTRACT FROM AUTHOR]

Cronobacter sakazakii is an opportunistic pathogen capable of causing severe infections, particularly in neonates. Despite the bacterium’s strong pathogenicity, the pathogenicity of C. sakazakii is not yet well understood. Using a comparative proteomic profiling approach, we successfully identified pdxY, encoding a pyridoxal kinase involved in the recycling of pyridoxal 59-phosphate (PLP), as a gene essential for the successful pathogenesis of C. sakazakii. Knocking out the pdxY gene resulted in slower growth and reduced virulence. Our study sheds light on the fundamental importance of pyridoxal kinase for the survival and virulence of C. sakazakii. The identification of pdxY as gene essential for successful pathogenesis provides a potential target for the development of new antibiotic treatments. IMPORTANCE The opportunistic pathogen Cronobacter sakazakii is known to cause severe infections, particularly in neonates, and can result in high mortality rates. In this study, we used a comparative proteomic profiling approach to identify genes essential for the successful pathogenesis of C. sakazakii. We successfully identified pdxY, encoding a pyridoxal kinase involved in the salvage pathway of pyridoxal 59-phosphate (PLP), as a gene essential for the successful pathogenesis of C. sakazakii. Knocking out the pdxY gene resulted in impaired growth and reduced virulence. This study sheds light on the fundamental importance of pyridoxal kinase for the survival and virulence of C. sakazakii, which can be a potential target for the development of new antibiotic treatments. This study highlights the importance of comparative proteomic profiling in identifying virulence factors that can be targeted for the development of new antibiotics. [ABSTRACT FROM AUTHOR]

(Macro)autophagy is a major lysosome-dependent degradation mechanism which engulfs, removes and recycles unwanted cytoplasmic material, including damaged organelles and toxic protein aggregates. Although a few studies implicate autophagy in CNS demyelinating pathologies, its role, particularly in mature oligodendrocytes and CNS myelin, remains poorly studied. Here, using both pharmacological and genetic inhibition of the autophagic machinery, we provide evidence that autophagy is an es-sential mechanism for oligodendrocyte maturation in vitro. Our study reveals that two core myelin proteins, namely proteolipid protein (PLP) and myelin basic protein (MBP) are incorporated into autophagosomes in oligodendrocytes, resulting in their degradation. Furthermore, we ablated atg5, a core gene of the autophagic machinery, specifically in myelinating glial cells in vivo by tamoxifen administration (plp-CreERT2; atg5 f/f) and showed that myelin maintenance is perturbed, leading to PLP accumulation. Significant morphological defects in myelin membrane such as de-compaction accompanied with increased axonal degeneration are observed. As a result, the mice exhibit behavioral deficits. In summary, our data highlight that the maintenance of adult myelin homeostasis in the CNS requires the involvement of a fully functional autophagic machinery.

The serine peptidase CLPP is conserved among bacteria, chloroplasts, and mitochondria. In humans and mice, its loss causes Perrault syndrome, which presents with growth deficits, infertility, deafness, and ataxia. In the filamentous fungus Podospora anserina, CLPP loss leads to longevity. CLPP substrates are selected by CLPX, an AAA+ unfoldase. CLPX is known to target delta-aminolevulinic acid synthase (ALAS) to promote pyridoxal phosphate (PLP) binding. CLPX may also influence cofactor association with other enzymes. Here, the evaluation of P. anserina metabolomics highlighted a reduction in arginine/histidine levels. In Mus musculus cerebellum, reductions in arginine/histidine and citrulline occurred with a concomitant accumulation of the heme precursor protoporphyrin IX. This suggests that the increased biosynthesis of 5-carbon (C5) chain deltaALA consumes not only C4 succinyl-CoA and C1 glycine but also specific C5 delta amino acids. As enzymes responsible for these effects, the elevated abundance of CLPX and ALAS is paralleled by increased OAT (PLP-dependent, ornithine delta-aminotransferase) levels. Possibly as a consequence of altered C1 metabolism, the proteome profiles of P. anserina CLPP-null cells showed strong accumulation of a methyltransferase and two mitoribosomal large subunit factors. The reduced histidine levels may explain the previously observed metal interaction problems. As the main nitrogen-storing metabolite, a deficiency in arginine would affect the urea cycle and polyamine synthesis. Supplementation of arginine and histidine might rescue the growth deficits of CLPP-mutant patients.

Thorough study of composition and fluorescence properties of a commercial reagent of active equine NAD-dependent alcohol dehydrogenase expressed and purified from E. coli has been carried out. Several experimental methods: spectral- and time-resolved two-photon excited fluorescence, sodium dodecyl sulfate–polyacrylamide gel electrophoresis, fast protein liquid chromatography, and mass spectrometry were used for analysis. The reagent under study was found to contain also a number of natural fluorophores: free NAD(P)H, NADH-alcohol dehydrogenase, NADPH-isocitrate dehydrogenase, and pyridoxal 5-phosphate—serine hydroxymethyltransferase complexes. The results obtained demonstrated the potential and limitations of popular optical methods as FLIM for separation of fluorescence signals from free and protein-bound forms of NADH, NADPH, and FAD that are essential coenzymes in redox reactions in all living cells. In particular, NADH-alcohol dehydrogenase and NADPH-isocitrate dehydrogenase complexes could not be optically separated in our experimental conditions although fast protein liquid chromatography and mass spectrometry analysis undoubtedly indicated the presence of both enzymes in the molecular sample used. Also, the results of fluorescence, fast protein liquid chromatography, and mass spectrometry analysis revealed a significant contribution of the enzyme-bound coenzyme pyridoxal 5-phosphate to the fluorescence signal that could be separated from enzyme-bound NADH by using bandpass filters, but could effectively mask contribution from enzyme-bound FAD because the fluorescence spectra of the species practically overlapped. It was shown that enzyme-bound pyridoxal 5-phosphate fluorescence can be separated from enzyme-bound NAD(P)H and FAD through analysis of short fluorescence decay times of about tens of picoseconds. However, this analysis was found to be effective only at relatively high number of peak photon counts in recorded fluorescence signals. The results obtained in this study can be used for interpretation of fluorescence signals from a mixture of enzyme-bound fluorophores and should be taken into consideration when determining the intracellular NADH/FAD ratio using FLIM. [ABSTRACT FROM AUTHOR]

Pyridoxal 5'-phosphate (PLP) is the main form of vitamin B6 in animal tissue and functions as a coenzyme for more than 160 different enzymatic reactions in the metabolism of amino acids, carbohydrates, lipids, and neurotransmitters. Estimated dietary intake of vitamin B6 and plasma PLP values differ a lot between studies, something which may be due to variable use of supplements, variations in dietary assessment and analytical methods. These factors make it difficult to achieve precise data for setting a correct recommended intake of vitamin B6. In addition, a plasma PLP concentration of 30 nmol/L is considered to be sufficient and the current recommendations for vitamin B6 intake is based on this concept. However, the metabolic marker for vitamin B6 status, HK ratio (HKr), starts to increase already when plasma PLP falls below 100 nmol/L and increases more steeply below 50 nmol/L, indicating biochemical deficiency. Consequently, a plasma PLP concentration of 30 nmol/L, may be too low as a marker for an adequate vitamin B6 status. [ABSTRACT FROM AUTHOR]

The excess accumulation of nonessential Al3+ in the human body can cause serious health problems, thus there is significant interest in the detection of Al3+. A new ratio fluorescent probe (RF‐probe) based on the mixing of red‐emitting D‐penicillamine (DPA) stabilized copper nanoclusters (DPA‐CuNCs) with pyridoxal 5'‐phosphate (PLP) was designed for sensitive and selective determination of Al3+. The −CHO group of PLP is expected to form an imine linkage with the free −NH2 group available of the DPA on the surface of DPA‐CuNCs. The content of Al3+ could be mensurated by the as‐prepared RF‐probe with a linear response in the range of 1.33–46.67 μmol ⋅ L−1, and corresponding limit of detection (LOD) was calculated to be 0.33 μmol ⋅ L−1. Moreover, the RF‐probe was successfully extended to the assay of Al3+ in aluminum carbonate chewing tablets, baking powder, pond water and drink water with satisfactory results compared to the standard atomic absorption spectroscopy (AAS). [ABSTRACT FROM AUTHOR]

Cofactor flavin adenine dinucleotide (FAD), a compound with flavin moiety and a derivative of riboflavin (vitamin B2), is shown to bind to Sox9 (a key transcription factor in early pancreatic development) and, subsequently, induce a large increase in markers of pancreatic development, including Ngn3 and PTF1a. Pyridoxal 5′-phosphate (PLP), the active form of vitamin B6, also binds to Sox9 and results in a similar increase in pancreatic development markers. Sox9 is known to be specifically important for pancreatic progenitors. Previously, there was no known link between FAD, PLP, or other co-factors and Sox9 for function. Thus, our findings show the mechanism by which FAD and PLP interact with Sox9 and result in the altered expression of pancreatic progenitor transcription factors involved in the pancreas development. [ABSTRACT FROM AUTHOR]

The current generation of sequencing technologies has led to significant advances in identifying novel disease-associated mutations and generated large amounts of data in a high-throughput manner. Such data in conjunction with clinical routine data are proven to be highly useful in deriving population-level and patient-level predictions, especially in the field of cancer precision medicine. However, data harmonization across multiple national and international clinical sites is an essential step for the assessment of events and outcomes associated with patients, which is currently not adequately addressed. The Observational Medical Outcomes Partnership (OMOP) Common Data Model (CDM) is an internationally established research data repository introduced by the Observational Health Data Science and Informatics (OHDSI) community to overcome this issue. To address the needs of cancer research, the genomic vocabulary extension was introduced in 2020 to support the standardization of subsequent data analysis. In this review, we evaluate the current potential of the OMOP CDM to be applicable in cancer prediction and how comprehensively the genomic vocabulary extension of the OMOP can serve current needs of AI-based predictions. For this, we systematically screened the literature for articles that use the OMOP CDM in predictive analyses in cancer and investigated the underlying predictive models/tools. Interestingly, we found 248 articles, of which most use the OMOP for harmonizing their data, but only 5 make use of predictive algorithms on OMOP-based data and fulfill our criteria. The studies present multicentric investigations, in which the OMOP played an essential role in discovering and optimizing machine learning (ML)-based models. Ultimately, the use of the OMOP CDM leads to standardized data-driven studies for multiple clinical sites and enables a more solid basis utilizing, e.g., ML models that can be reused and combined in early prediction, diagnosis, and improvement of personalized cancer care and biomarker discovery. [ABSTRACT FROM AUTHOR]

The serine hydroxymethyltransferase (SHMT; E.C. 2.1.2.1) is involved in the interconversion of serine/glycine and tetrahydrofolate (THF)/5,10-methylene THF, playing a key role in one-carbon metabolism, the de novo purine pathway, cellular methylation reactions, redox homeostasis maintenance, and methionine and thymidylate synthesis. GmSHMT08 is the soybean gene underlying soybean cyst nematode (SCN) resistance at the Rhg4 locus. GmSHMT08 protein contains four tetrahydrofolate (THF) cofactor binding sites (L129, L135, F284, N374) and six pyridoxal phosphate (PLP) cofactor binding/catalysis sites (Y59, G106, G107, H134, S190A, H218). In the current study, proteomic analysis of a data set of protein complex immunoprecipitated using GmSHMT08 antibodies under SCN infected soybean roots reveals the presence of enriched pathways that mainly use glycine/serine as a substrate (glyoxylate cycle, redox homeostasis, glycolysis, and heme biosynthesis). Root and leaf transcriptomic analysis of differentially expressed genes under SCN infection supported the proteomic data, pointing directly to the involvement of the interconversion reaction carried out by the serine hydroxymethyltransferase enzyme. Direct site mutagenesis revealed that all mutated THF and PLP sites at the GmSHMT08 resulted in increased SCN resistance. We have shown the involvement of PLP sites in SCN resistance. Specially, the effect of the two Y59 and S190 PLP sites was more drastic than the tested THF sites. This unprecedented finding will help us to identify the biological outcomes of THF and PLP residues at the GmSHMT08 and to understand SCN resistance mechanisms. [ABSTRACT FROM AUTHOR]

Pyridoxal 5′-phosphate (PLP) is the active form of vitamin B6, but it is highly reactive and poisonous in its free form. YggS is a PLP-binding protein found in bacteria and humans that mediates PLP homeostasis by delivering PLP to target enzymes or by performing a protective function. Several biochemical and structural studies of YggS have been reported, but the mechanism by which YggS recognizes PLP has not been fully elucidated. Here, we report a functional and structural analysis of YggS from Fusobacterium nucleatum (FnYggS). The PLP molecule could bind to native FnYggS, but no PLP binding was observed for selenomethionine (SeMet)-derivatized FnYggS. The crystal structure of FnYggS showed a type III TIM barrel fold, exhibiting structural homology with several other PLP-dependent enzymes. Although FnYggS exhibited low (<35%) amino acid sequence similarity with previously studied YggS proteins, its overall structure and PLP-binding site were highly conserved. In the PLP-binding site of FnYggS, the sulfate ion was coordinated by the conserved residues Ser201, Gly218, and Thr219, which were positioned to provide the binding moiety for the phosphate group of PLP. The mutagenesis study showed that the conserved Ser201 residue in FnYggS was the key residue for PLP binding. These results will expand the knowledge of the molecular properties and function of the YggS family. [ABSTRACT FROM AUTHOR]

Background Folate-mediated 1-carbon metabolism requires several nutrients, including vitamin B6. Circulating biomarker concentrations indicating high vitamin B6 status are associated with a reduced risk of colorectal cancer (CRC). However, little is known about the effect of B6 status in relation to clinical outcomes in CRC patients. Objectives We investigated survival outcomes in relation to vitamin B6 status in prospectively followed CRC patients. Methods A total of 2031 patients with stage I–III CRC participated in 6 prospective patient cohorts in the international FOCUS (folate-dependent 1-carbon metabolism in colorectal cancer recurrence and survival) Consortium. Preoperative blood samples were used to measure vitamin B6 status by the direct marker pyridoxal 5′-phosphate (PLP), as well as the functional marker HK-ratio (HKr)[3′-hydroxykynurenine: (kynurenic acid + xanthurenic acid + 3′-hydroxy anthranilic acid + anthranilic acid)]. Using Cox proportional hazards regression, we examined associations of vitamin B6 status with overall survival (OS), disease-free survival (DFS), and risk of recurrence, adjusted for patient age, sex, circulating creatinine concentrations, tumor site, stage, and cohort. Results After a median follow-up of 3.2 y for OS, higher preoperative vitamin B6 status as assessed by PLP and the functional marker HKr was associated with 16–32% higher all-cause and disease-free survival, although there was no significant association with disease recurrence (doubling in PLP concentration: HROS, 0.68; 95% CI: 0.59, 0.79; HRDFS, 0.84; 95% CI: 0.75, 0.94; HRRecurrence, 0.96; 95% CI: 0.84, 1.09; HKr: HROS, 2.04; 95% CI: 1.67, 2.49; HRDFS, 1.56; 95% CI: 1.31, 1.85; HRRecurrence, 1.21; 95% CI: 0.96,1. 52). The association of PLP with improved OS was consistent across colorectal tumor site (right-sided colon: HROS, 0.75; 95% CI: 0.59, 0.96; left-sided colon: HROS, 0.71; 95% CI: 0.55, 0.92; rectosigmoid junction and rectum: HROS, 0.61; 95% CI: 0.47, 0.78). Conclusion Higher preoperative vitamin B6 status is associated with improved OS among stage I–III CRC patients. [ABSTRACT FROM AUTHOR]

Radical aminomutases are pyridoxal 5′-phosphate (PLP, a B6 vitamer)-dependent enzymes that require the generation of a 5′-deoxyadenosyl radical to initiate the catalytic cycle, to perform a 1,2 amino group shift reaction. The role of the nitrogen atom of PLP in radical aminomutases has not been investigated extensively yet. We report an alternative synthetic procedure to provide easy access to 1-deazaPLP (dAPLP), an isosteric analog of PLP which acts as a probe for studying the role of the nitrogen atom. Our results revealed that lysine 5,6-aminomutase (5,6-LAM), a radical aminomutase, reconstituted with dAPLP cannot turn over a substrate, demonstrating that the nitrogen atom is essential for radical aminomutases. In contrast, biochemical and spectroscopic studies on the S238A variant reconstituted with PLP revealed a minuscule loss of activity. This apparent anomaly can be explained by a water-mediated rescue of activity in S238A, as if mimicking the active site of lysine 2,3-aminomutase. This study leads to a better comprehension of how enzymes harness the optimum capability of PLP to realize catalysis. [ABSTRACT FROM AUTHOR]

Effective biocatalysts for the synthesis of optically pure amines from keto precursors are highly required in organic synthesis. Transaminases are a large group of PLP-dependent enzymes, which can be utilized for production of chiral amines or amino acids. The bioinformatic approach previously made to search for promising transaminases with unusual characteristics surprisingly revealed mysterious genes in some Gram-negative bacteria, which products were annotated as aminotransferases, but they lacked the key catalytic lysine residue required for covalent binding of the PLP-cofactor. To address the question of which products these genes encode, we obtained the first structure of such a type of protein from the bacterium Variovorax paradoxus (VP5454) and provided its comprehensive analysis. We demonstrated that VP5454 has a typical aminotransferase fold and architecture of the active site, where substitution of the catalytic lysine with asparagine was observed. Despite that no covalent adduct can be formed between PLP and asparagine residue, using X-ray analysis and molecular dynamic (MD) simulation, we demonstrated that VP5454 is able to bind the PLP molecule in the transaminase in a specific manner, with PLP coordinated via its phosphate moiety. Taking into account a number of sequences homologous to VP5454 with a substituted catalytic lysine found in the genomes of various bacteria, we speculate that the proteins encoded by these sequences may have hidden functional roles. [ABSTRACT FROM AUTHOR]

This paper seeks to detect Parkinson's disease among healthy cases, several neurological diseases, in particular those which are very similar, that is to say representing a parkinsonian syndrome such as multi system atrophy. Early detection based on the phonatory symptoms will offer a possibility to the treatments proposed by the doctors to act effectively on the patient hence the interest of our project. We used several algorithms such as MFCC, PLP and RASTA PLP in addition to our new technique called Real Cepstral Discrete Cosine Transform with an SVM classifier with its different kernels to guarantee a better detection. [ABSTRACT FROM AUTHOR]

Background: Detecting cortical demyelination using magnetic resonance imaging (MRI) in multiple sclerosis (MS) remains a challenge. Magnetization transfer ratio (MTR), T1-weighted/T2-weighted ratio (T1T2R), and T2-weighted (T2w) signal are sensitive to cortical demyelination, but their accuracy is unknown. Objectives: To quantify the sensitivity, specificity, and accuracy of postmortem T1T2R, MTR, and T2w in detecting cortical demyelination. Methods: In situ postmortem MRIs from 9 patients were used to measure T1T2R, MTR, and T2w along the midline of cortical gray matter and classified as normal or abnormal. MRIs were co-registered and compared to hemispheric myelin staining. The sensitivity, specificity, and accuracy of T1T2R, MTR, and T2w in detecting cortical demyelination were measured. Results: The mean age (standard deviation) at death was 64.7 (+/-13.7) years with a disease duration of 23.8 (+/-10.5) years. The sensitivity was 78% for MTR, 75% for T1T2R, and 63% for T2w. The specificity was 46% (T2w), 13% (T1T2R), and 29% (MTR). The accuracy was 71% (T2w), 39% (MTR), and 42% (T1T2R). There were no significant differences between different MRI measures in cortical demyelination or intracortical/subpial lesion detection. Conclusions: Although somewhat sensitive, the modest specificity of conventional MRI modalities for cortical demyelination indicates that they are influenced by cortical changes other than demyelination. Improved acquisition and post-processing are needed to reliably measure cortical lesion load. [ABSTRACT FROM AUTHOR]

[Display omitted] • A novel concept of demonstrated pyridoxal phosphate permeability in E.coli • Pyridoxal phosphate inclusion in media increases protein yield and enzymatic activity • A novel protein model system for pyridoxal phosphate linked permeation studies. PLP (Pyridoxal 5′-phosphate) dependent proteins are important drug targets and effector molecules, and thus, their heterologous overexpression is of industrial importance and has commercial value. We have predicted the docking site of PLP on O-acetylserine sulphdrylase (OASS) of H.contortus and determined that the lysine-47 is very important for the binding of PLP. We have used this protein as a model protein for testing the effect of PLP on the expression of PLP dependent proteins by E.coli. Soluble recombinant protein could be purified from each of the culture vials grown with variable amount of PLP [0 mM (Group I), 0.01 mM (Group II), 0.025 mM (Group III), 0.05 mM (Group IV) and 0.1 mM (Group V)]. There was approximately 4.2 %, 7.2 %, 10.5 % and 18 % increase in purified protein yield in Group II, III, IV and V, respectively, in comparison to group I. There was a significant quenching of tryptophan fluorescence emission in groups II, III, IV and V compared to group I. We could find a PLP concentration-dependent increase in expression and catalytic activity signifying the probable transport of PLP across the membrane. E.coli does not secrete any dephosphorylation factor for PLP in the immediate microenvironment. There was a significant reduction in levels of inorganic phosphate after PLP dephosphorylation in induced group in comparison to uninduced E.coli and BL21 groups, signifying PLP internalization in E.coli. The mechanism of transport of PLP in the light of the current experiment is still unknown and should be a subject of future studies. [ABSTRACT FROM AUTHOR]

Enzymatic amino acid assays are important in physiological research and clinical diagnostics because abnormal amino acid concentrations in biofluids are associated with various diseases. L-histidine decarboxylase from Photobacterium phosphoreum (PpHDC) is a pyridoxal 5′-phosphate-dependent enzyme and a candidate for use in an L-histidine quantitation assay. Previous cysteine substitution experiments demonstrated that the PpHDC C57S mutant displayed improved long-term storage stability and thermostability when compared with those of the wild-type enzyme. In this study, combinational mutation experiments of single cysteine substitution mutants of PpHDC were performed, revealing that the PpHDC C57S/C101V/C282V mutant possessed the highest thermostability. The stabilizing mechanism of these mutations was elucidated by solving the structures of PpHDC C57S and C57S/C101V/C282V mutants by X-ray crystallography. In the crystal structures, two symmetry-related PpHDC molecules form a domain-swapped homodimer. The side chain of S57 is solvent exposed in the structure, indicating that the C57S mutation eliminates chemical oxidation or disulfide bond formation with a free thiol group, thereby providing greater stability. Residues 101 and 282 form hydrophobic interactions with neighboring hydrophobic residues. Mutations C101V and C282V enhanced thermostability of PpHDC by filling a cavity present in the hydrophobic core (C101V) and increasing hydrophobic interactions. [ABSTRACT FROM AUTHOR]

The compatible solutes ectoine and 5-hydroxyectoine are widely synthesized by bacteria as osmostress protectants. These nitrogen-rich tetrahydropyrimidines can also be exploited as nutrients by microorganisms. Many ectoine/5-hydroxyectoine catabolic gene clusters are associated with a regulatory gene (enuR : ectoine nutrient utilization regulator) encoding a repressor protein belonging to the MocR/GabR sub-family of GntR-type transcription factors. Focusing on EnuR from the marine bacterium Ruegeria pomeroyi , we show that the dimerization of EnuR is mediated by its aminotransferase domain. This domain can fold independently from its amino-terminal DNA reading head and can incorporate pyridoxal-5′-phosphate (PLP) as cofactor. The covalent attachment of PLP to residue Lys302 of EnuR was proven by mass-spectrometry. PLP interacts with system-specific, ectoine and 5-hydroxyectoine-derived inducers: alpha-acetyldiaminobutyric acid (alpha-ADABA), and hydroxy-alpha-acetyldiaminobutyric acid (hydroxy-alpha-ADABA), respectively. These inducers are generated in cells actively growing with ectoines as sole carbon and nitrogen sources, by the EutD hydrolase and targeted metabolic analysis allowed their detection. EnuR binds these effector molecules with affinities in the low micro-molar range. Studies addressing the evolutionary conservation of EnuR, modelling of the EnuR structure, and docking experiments with the inducers provide an initial view into the cofactor and effector binding cavity. In this cavity, the two high-affinity inducers for EnuR, alpha-ADABA and hydroxy-alpha-ADABA, are positioned such that their respective primary nitrogen group can chemically interact with PLP. Purified EnuR bound with micro-molar affinity to a 48 base pair DNA fragment containing the sigma-70 type substrate-inducible promoter for the ectoine/5-hydroxyectoine importer and catabolic gene cluster. Consistent with the function of EnuR as a repressor, the core elements of the promoter overlap with two predicted EnuR operators. Our data lend themselves to a straightforward regulatory model for the initial encounter of EnuR-possessing ectoine/5-hydroxyectoine consumers with environmental ectoines and for the situation when the external supply of these compounds has been exhausted by catabolism. [ABSTRACT FROM AUTHOR]

This paper implements the continuous Hindi Automatic Speech Recognition (ASR) system using the proposed integrated features vector with Recurrent Neural Network (RNN) based Language Modeling (LM). The proposed system also implements the speaker adaptation using Maximum-Likelihood Linear Regression (MLLR) and Constrained Maximum likelihood Linear Regression (C-MLLR). This system is discriminatively trained by Maximum Mutual Information (MMI) and Minimum Phone Error (MPE) techniques with 256 Gaussian mixture per Hidden Markov Model(HMM) state. The training of the baseline system has been done using a phonetically rich Hindi dataset. The results show that discriminative training enhances the baseline system performance by up to 3%. Further improvement of ~7% has been recorded by applying RNN LM. The proposed Hindi ASR system shows significant performance improvement over other current state-of-the-art techniques.

Thorough study of composition and fluorescence properties of a commercial reagent of active equine NAD-dependent alcohol dehydrogenase expressed and purified from E. coli has been carried out. Several experimental methods: spectral- and time-resolved two-photon excited fluorescence, sodium dodecyl sulfate–polyacrylamide gel electrophoresis, fast protein liquid chromatography, and mass spectrometry were used for analysis. The reagent under study was found to contain also a number of natural fluorophores: free NAD(P)H, NADH-alcohol dehydrogenase, NADPH-isocitrate dehydrogenase, and pyridoxal 5-phosphate—serine hydroxymethyltransferase complexes. The results obtained demonstrated the potential and limitations of popular optical methods as FLIM for separation of fluorescence signals from free and protein-bound forms of NADH, NADPH, and FAD that are essential coenzymes in redox reactions in all living cells. In particular, NADH-alcohol dehydrogenase and NADPH-isocitrate dehydrogenase complexes could not be optically separated in our experimental conditions although fast protein liquid chromatography and mass spectrometry analysis undoubtedly indicated the presence of both enzymes in the molecular sample used. Also, the results of fluorescence, fast protein liquid chromatography, and mass spectrometry analysis revealed a significant contribution of the enzyme-bound coenzyme pyridoxal 5-phosphate to the fluorescence signal that could be separated from enzyme-bound NADH by using bandpass filters, but could effectively mask contribution from enzyme-bound FAD because the fluorescence spectra of the species practically overlapped. It was shown that enzyme-bound pyridoxal 5-phosphate fluorescence can be separated from enzyme-bound NAD(P)H and FAD through analysis of short fluorescence decay times of about tens of picoseconds. However, this analysis was found to be effective only at relatively high number of peak photon counts in recorded fluorescence signals. The results obtained in this study can be used for interpretation of fluorescence signals from a mixture of enzyme-bound fluorophores and should be taken into consideration when determining the intracellular NADH/FAD ratio using FLIM.

The compatible solutes ectoine and 5-hydroxyectoine are widely synthesized by bacteria as osmostress protectants. These nitrogen-rich tetrahydropyrimidines can also be exploited as nutrients by microorganisms. Many ectoine/5-hydroxyectoine catabolic gene clusters are associated with a regulatory gene (enuR: ectoine nutrient utilization regulator) encoding a repressor protein belonging to the MocR/GabR sub-family of GntR-type transcription factors. Focusing on EnuR from the marine bacterium Ruegeria pomeroyi, we show that the dimerization of EnuR is mediated by its aminotransferase domain. This domain can fold independently from its amino-terminal DNA reading head and can incorporate pyridoxal-5′-phosphate (PLP) as cofactor. The covalent attachment of PLP to residue Lys302 of EnuR was proven by mass-spectrometry. PLP interacts with system-specific, ectoine and 5-hydroxyectoine-derived inducers: alpha-acetyldiaminobutyric acid (alpha-ADABA), and hydroxy-alpha-acetyldiaminobutyric acid (hydroxy-alpha-ADABA), respectively. These inducers are generated in cells actively growing with ectoines as sole carbon and nitrogen sources, by the EutD hydrolase and targeted metabolic analysis allowed their detection. EnuR binds these effector molecules with affinities in the low micro-molar range. Studies addressing the evolutionary conservation of EnuR, modelling of the EnuR structure, and docking experiments with the inducers provide an initial view into the cofactor and effector binding cavity. In this cavity, the two high-affinity inducers for EnuR, alpha-ADABA and hydroxy-alpha-ADABA, are positioned such that their respective primary nitrogen group can chemically interact with PLP. Purified EnuR bound with micro-molar affinity to a 48 base pair DNA fragment containing the sigma-70 type substrate-inducible promoter for the ectoine/5-hydroxyectoine importer and catabolic gene cluster. Consistent with the function of EnuR as a repressor, the core elements of the promoter overlap with two predicted EnuR operators. Our data lend themselves to a straightforward regulatory model for the initial encounter of EnuR-possessing ectoine/5-hydroxyectoine consumers with environmental ectoines and for the situation when the external supply of these compounds has been exhausted by catabolism.

1,4-diaminobutane is widely used in the industrial production of polymers, pharmaceuticals, agrochemicals and surfactants. Owing to economic and environmental concerns, there has been a growing interest in using microbes to produce 1,4-diaminobutane. However, there is lack of research on the influence of cofactors pyridoxal phosphate (PLP) and NADPH on the synthesis of 1,4-diaminobutane. PLP serves as a cofactor of ornithine decarboxylase in the synthesis of 1,4-diaminobutane. Additionally, the synthesis of 1 mol 1,4-diaminobutane requires 2 mol NADPH, thus necessitating consideration of NADPH balance in the efficient synthesis of 1,4-diaminobutane by Escherichia coli . The aim of this study was to enhance the synthesis efficiency of 1,4-diaminobutane through increasing production of PLP and NADPH. By optimizing the expression of the genes associated with synthesis of PLP and NADPH in E. coli , cellular PLP and NADPH levels increased, and the yield of 1,4-diaminobutane also increased accordingly. Ultimately, using glucose as the primary carbon source, the yield of 1,4-diaminobutane in the recombinant strain NAP19 reached 272 mg/L·DCW, by increased 79% compared with its chassis strain.