Decarboxylase Enzyme Research Articles

Functional and technological characterization of lactic acid bacteria isolated from Turkish dry-fermented sausage (sucuk).

In this study, 10 lactic acid bacteria were isolated from Turkish fermented sausage (sucuk) and identified as 5 Lactobacillus plantarum, 1 Pediococcus acidilactici, 1 Weissella hellenica, 1 Lactobacillus pentosus, and 2 Lactobacillus sakei. PCR screening of genes encoding plantaricin A and pediocin showed the presence of plantaricin A gene in 9 and pediocin gene in 3 of strains. All isolates showed antibacterial and antifungal effect on most of the tested microorganisms. gad gene, encoding glutamic acid decarboxylase enzyme, was detected in all isolates except Weisella hellenica KS-24. Eight of isolates were determined as gamma-amino butyric acid (GABA) producer in the presence of 53mM mono sodium glutamate (MSG) by HPLC and TLC analysis. DPPH scavenging activity was observed for all isolates. Additionally, isolates were able to produce exopolysaccharide in the presence of sucrose. The best exopolysaccharide (EPS) production was achieved with L. plantarum KS-11 and L. pentosus KS-27. As a result, this study characterized some techno-functional properties of LAB isolates from sucuk. It was concluded that the isolates studied have the potential to be used in obtaining functional products in meat industry, as well as strain selection may be effective in providing the desired properties in the product.

Parkinson's Disease Medication Alters Small Intestinal Motility and Microbiota Composition in Healthy Rats.

ABSTRACTParkinson’s disease (PD) is known to be associated with altered gastrointestinal function and microbiota composition. To date, the effect of PD medication on the gastrointestinal function and microbiota, at the site of drug absorption, the small intestine, has not been studied, although it may represent an important confounder in reported microbiota alterations observed in PD patients. To this end, healthy (non-PD) wild-type Groningen rats were employed and treated with dopamine, pramipexole (in combination with levodopa-carbidopa), or ropinirole (in combination with levodopa-carbidopa) for 14 sequential days. Rats treated with dopamine agonists showed a significant reduction in small intestinal motility and an increase in bacterial overgrowth in the distal small intestine. Notably, significant alterations in microbial taxa were observed between the treated and vehicle groups; analogous to the changes previously reported in human PD versus healthy control microbiota studies. These microbial changes included an increase in Lactobacillus and Bifidobacterium and a decrease in Lachnospiraceae and Prevotellaceae. Markedly, certain Lactobacillus species correlated negatively with levodopa levels in the systemic circulation, potentially affecting the bioavailability of levodopa. Overall, the study highlights a significant effect of PD medication intrinsically on disease-associated comorbidities, including gastrointestinal dysfunction and small intestinal bacterial overgrowth, as well as the gut microbiota composition. The results urge future studies to take into account the influence of PD medication per se when seeking to identify microbiota-related biomarkers for PD.IMPORTANCE Parkinson’s disease (PD) is the second most common neurodegenerative disorder and is known to be associated with altered gastrointestinal function and microbiota composition. We previously showed that the gut bacteria harboring tyrosine decarboxylase enzymes interfere with levodopa, the main treatment for PD (S. P. van Kessel, A. K. Frye, A. O. El-Gendy, M. Castejon, A. Keshavarzian, G. van Dijk, and S. El Aidy, Nat Commun 10:310, 2019). Although PD medication could be an important confounder in the reported alterations, its effect, apart from the disease itself, on the microbiota composition or the gastrointestinal function at the site of drug absorption, the small intestine, has not been studied. The findings presented here show a significant impact of commonly prescribed PD medication on the small intestinal motility, small intestinal bacterial overgrowth, and microbiota composition, irrespective of the PD. Remarkably, we observed negative associations between bacterial species harboring tyrosine decarboxylase activity and levodopa levels in the systemic circulation, potentially affecting the bioavailability of levodopa. Overall, this study shows that PD medication is an important factor in determining gastrointestinal motility and, in turn, microbiota composition and may, partly, explain the differential abundant taxa previously reported in the cross-sectional PD microbiota human studies. The results urge future studies to take into account the influence of PD medication on gut motility and microbiota composition when seeking to identify microbiota-related biomarkers for PD.

In-Vivo and In-Silico Evidence of Antitrypanocidal Activities of Selected Plants from Asteraceae Family against Trypanosoma brucei brucei

The menace of animal African trypanosomiasis (AAT) has increased in sub-Saharan Africa despite countless interventions through chemotherapy and health policies. The study evaluated the in-vivo antitrypanosomal activities of five plant species from the Asteraceae family and further subjected some previously isolated compounds from the active plants to molecular docking on an identified T. brucei target. The extracts of the aerial parts of Tithonia diversifolia, Chromolaena odorata, Aspilia africana, Vernonia glaberrima, and Synedrella nodiflora were subjected to in-vivo antitrypanosomal activity against T. b. brucei-infected mice by monitoring the parasitemia and packed cell volume (PCV) at intervals. Some isolated compounds were docked to a co-crystallized ornithine decarboxylase (ODC) enzyme target, and the binding energies were compared. All the extracts (200-600 mg/kg) caused a significant (p < 0.05) decline in the parasitemia level compared with the untreated group. Both T. diversifolia (200-600 mg/kg) and V. glaberrima (400-600 mg/kg) caused total clearance of the parasite from the bloodstream of infected mice by the 17th day. Similarly, except for S. nodiflora and C. odorata, all other tested extracts restored the PCV of the infected mice to the baseline value (> 40 %). Woodhousin from T. diversifolia and apigenin from V. glaberrima interacted maximally with the ODC of T. b. brucei. The study has established the basis for using the selected plants in the management of AAT in folk medicine and further provided insights into the potential of the plant species to provide new bioactive lead compounds.

Immunocytochemical evidence of histamine 1 and histamine 2 receptors on mice sperm

Histamine is a biogenic amine which is synthesised by L-histidine decarboxylase enzyme (HDC). The histamine 1 and 2 antagonist administrations have been highly reported to cause detrimental effect on sperm parameters, which arisen the speculation of histamine 1 (H1R) and histamine 2 (H2R) receptors might be present in sperm. The present study was aimed to provide evidence on the localisation of H1R and H2R on mice sperm through immunocytochemistry. The sperm was harvested from cauda epididymis. After one hour of incubation, sperm suspension was smeared onto a poly-lysine-coated slide and allowed to dry before fixation and permeabilisation processes. The primary antibody encoded for receptors was exposed to the fluorescently tagged antibody; fluorescein isothiocyanate (FITC) conjugate followed by nuclear staining with 4, 6-diamino-2-phenylindole dihydrochloride (DAPI). The testis, stomach, and skin were used as the positive controls. Our data showed that both receptors have been expressed on the midpiece and acrosome of mice. The present result was the first discovery of the presence and immunolocalisation of H1R and H2R on mice sperm. Therefore, present study proposes that these receptors could be involved in calcium regulatory mechanism and protein phosphorylation which are responsible for fertilisation-related processes.

Postharvest γ-aminobutyric acid application mitigates chilling injury of aonla (Emblica officinalis Gaertn.) fruit during low temperature storage

Aonla is a nutrients rich underutilized subtropical fruit. It has short shelf life and is prone to chilling injury (CI) below 6 °C. So, the effect of γ-aminobutyric acid [5 mmol L−1 (GABA)] was investigated on aonla fruit CI mitigation during low temperature storage at 5 ± 1 °C for 24 d. Results showed that GABA treatment markedly suppressed CI, decay incidence, membrane permeability, malondialdehyde, superoxide anion and hydrogen peroxide in contrast with control. Treated fruit showed higher activity of glutamate decarboxylase (GAD) and GABA transaminase (GABA-T) enzymes which ultimately resulted in increased endogenous GABA content accumulation. Similarly, GABA application stimulated the activities of Δ1-pyrroline-5-carboxylate synthetase (P5CS) and ornithine aminotransferase (OAT) enzymes which thus enhanced proline accumulation. The GABA treated fruit exhibited higher phenylalanine ammonia lyase (PAL) and reduced polyphenol oxidase (PPO) enzyme activity and showed higher total phenols accumulation. In addition, total flavonoids, ascorbic acid and glutathione content were also significantly higher in GABA treated aonla fruit. In addition, GABA treated aonla fruit exhibited markedly higher ascorbate peroxidase (APX) catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD) activities in comparison with the control. In addition, GABA treated fruit showed lower total soluble solids, ripening index and juice pH along with markedly higher concentration of titratable acidity. In conclusion, GABA treatment could be used for CI mitigation and overall quality maintenance of cold stored aonla fruit.

ER-localized phosphatidylethanolamine synthase plays a conserved role in lipid droplet formation.

The asymmetric distribution of phospholipids in membranes is a fundamental principle of cellular compartmentalization and organization. Phosphatidylethanolamine (PE), a nonbilayer phospholipid that contributes to organelle shape and function, is synthesized at several subcellular localizations via semiredundant pathways. Previously, we demonstrated in budding yeast that the PE synthase Psd1, which primarily operates on the mitochondrial inner membrane, is additionally targeted to the ER. While ER-localized Psd1 is required to support cellular growth in the absence of redundant pathways, its physiological function is unclear. We now demonstrate that ER-localized Psd1 sublocalizes on the ER to lipid droplet (LD) attachment sites and show it is specifically required for normal LD formation. We also find that the role of phosphatidylserine decarboxylase (PSD) enzymes in LD formation is conserved in other organisms. Thus we have identified PSD enzymes as novel regulators of LDs and demonstrate that both mitochondria and LDs in yeast are organized and shaped by the spatial positioning of a single PE synthesis enzyme.

Computational analysis of non-synonymous single nucleotide polymorphism in UROD gene linked with familial Porphyria Cutanea Tarda

Porphyria Cutanea Tarda (PCT) is the most common type of human porphyria, characterized by onycholysis and blisters on the skin. It is a multifactorial disorder, mainly characterized by reduced uroporphyrinogen decarboxylase enzyme activity. Non-synonymous single nucleotide polymorphisms (nsSNPs) in UROD gene resulted in amino acid sequence alteration in protein product. Previous studies have reported that nsSNPs in UROD gene linked with familial PCT. The preliminary aim of current study is to evaluate the effect of nsSNPs on structure and function of UROD enzyme by developing the secondary and tertiary structures, computationally. Out of 294 nsSNPs, 115 nsSNPs within the coding region have been analyzed as deleterious by means of all evaluated tools, suggesting their involvement in downregulation of UROD gene expression and its protein product functioning. In future, the current study would be used to perform in-vitro analysis of reported nsSNPs, which ultimately lead towards the effective drug designing strategies.

Safety of AADC Gene Therapy for Moderately Advanced Parkinson Disease: Three-Year Outcomes From the PD-1101 Trial.

Background and ObjectivesTo report final, 36-month safety and clinical outcomes from the PD-1101 trial of NBIb-1817 (VY-AADC01) in participants with moderately advanced Parkinson disease (PD) and motor fluctuations.MethodsPD-1101 was a phase 1b, open-label, dose escalation trial of VY-AADC01, an experimental AAV2 gene therapy encoding the human aromatic l-amino acid decarboxylase (AADC) enzyme. VY-AADC01 was delivered via bilateral, intraoperative MRI-guided putaminal infusions to 3 cohorts (n = 5 participants per cohort): cohort 1, ≤7.5 × 1011 vector genomes (vg); cohort 2, ≤1.5 × 1012 vg; cohort 3, ≤4.7 × 1012 vg.ResultsNo serious adverse events (SAEs) attributed to VY-AADC01 were reported. All 4 non-vector–related SAEs (atrial fibrillation and pulmonary embolism in 1 participant and 2 events of small bowel obstruction in another participant) resolved. Requirements for PD medications were reduced by 21%–30% in the 2 highest dose cohorts at 36 months. Standard measures of motor function (PD diary, Unified Parkinson's Disease Rating Scale III “off”-medication and “on”-medication scores), global impressions of improvement (Clinical Global Impression of Improvement, Patient Global Impression of Improvement), and quality of life (39-item Parkinson's Disease Questionnaire) were stable or improved compared with baseline at 12, 24, and 36 months following VY-AADC01 administration across cohorts.DiscussionsVY-AADC01 and the surgical administration procedure were well-tolerated and resulted in stable or improved motor function and quality of life across cohorts, as well as reduced PD medication requirements in cohorts 2 and 3 over 3 years.Trial Registration InformationNCT01973543.Classification of EvidenceThis study provides Class IV evidence that, in patients with moderately advanced PD and motor fluctuations, putaminal infusion of VY-AADC01 is well tolerated and may improve motor function.

Clinical presentation, comorbidities and treatment of GAD antibodies associated stiff person syndrome

Stiff Person Syndrome (SPS) is a rare autoimmune neurological disorder, characterized by axial and limb muscular stiffness and spasms, due to a loss of physiological GABAergic CNS inhibition. In 60%–80% of patients with classic SPS phenotype antibodies against glutamic acid decarboxylase (GAD) enzyme can be detected.

Transcriptomic and Metabolomic Analyses Reveals That Exogenous Methyl Jasmonate Regulates Galanthamine Biosynthesis in Lycoris longituba Seedlings.

The Amaryllidaceae alkaloid galanthamine (Gal) in Lycoris longituba is a secondary metabolite that has been used to treat Alzheimer’s disease. Plant secondary metabolism is affected by methyl jasmonate (MeJA) exposure, although the regulatory mechanisms of MeJA on L. longituba seedlings remains largely unknown. In the present study, 75, 150, and 300 μM MeJA were used as treatments on L. longituba seedlings for 7, 14, 21, and 28 days, while 0 μM MeJA was used as the control (MJ-0). The effect of exogenous MeJA on Gal synthesis in L. longituba was then investigated using transcriptomic sequencing and metabolite profiling via GC-MS and LC-MS analysis. Galanthamine (Gal), lycorine (Lyc), and lycoramine (Lycm) abundances were 2. 71-, 2. 01-, and 2.85-fold higher in 75 μM MeJA (MJ-75) treatment plants compared to MJ-0 treatment plants after 7 days of cultivation. Transcriptomic analysis further showed that MJ-75 treatment significantly induced the expression of norbelladine synthase (NBS) and norbelladine 4′-O-methyltransferase (OMT), which are involved in the Gal biosynthesis pathway. In addition, increased expression was observed in MJ-75 treatment plants for genes in the JA synthesis and JA signaling pathways including those of allene oxide cyclase (AOC), 12-oxo-phytodienoic acid reductase (OPR), jasmonic acid amino acid synthase (JAR), and transcription factor MYC. The L. longituba tyrosine decarboxylase (LlTYDC) enzyme was identified and proposed to be involved in the Gal biosynthetic pathway. Metabolomics results demonstrated that the accumulation of Amaryllidaceae alkaloids, and especially alkaloids in the Gal biosynthesis pathway, could be induced by MJ-75 treatment. Interestingly, metabolites in the JA synthesis pathway were also affected by MeJA treatment. Overall, this multi-omics study suggests that both the JA synthesis/JA signaling and Gal biosynthesis pathways were affected by exogenous MeJA treatment. This comprehensive study of gene expression and metabolite contents can help us better understand the molecular mechanisms underlying MeJA-mediated Gal biosynthesis in L. longituba.

TDP-43 regulates GAD1 mRNA splicing and GABA signaling in Drosophila CNS

Alterations in the function of the RNA-binding protein TDP-43 are largely associated with the pathogenesis of amyotrophic lateral sclerosis (ALS), a devastating disease of the human motor system that leads to motoneurons degeneration and reduced life expectancy by molecular mechanisms not well known. In our previous work, we found that the expression levels of the glutamic acid decarboxylase enzyme (GAD1), responsible for converting glutamate to γ-aminobutyric acid (GABA), were downregulated in TBPH-null flies and motoneurons derived from ALS patients carrying mutations in TDP-43, suggesting that defects in the regulation of GAD1 may lead to neurodegeneration by affecting neurotransmitter balance. In this study, we observed that TBPH was required for the regulation of GAD1 pre-mRNA splicing and the levels of GABA in the Drosophila central nervous system (CNS). Interestingly, we discovered that pharmacological treatments aimed to potentiate GABA neurotransmission were able to revert locomotion deficiencies in TBPH-minus flies, revealing novel mechanisms and therapeutic strategies in ALS.

Long-term subculture affects rooting competence via changes in the hormones and protein profiles in Cedrela fissilis Vell. (Meliaceae) shoots

Long-term subculture plays an essential role in the large-scale multiplication and production of somatic plantlets. We investigated the effects of long-term subculture on in vitro shoot development and ex vitro rooting associated with changes in the hormones and protein profiles in Cedrela fissilis. The number of subcultures of shoots induced a decrease in the ex vitro rooting response. The reduction in adventitious root (AR) formation was associated with decreases in the contents of indole-3-acetic acid (IAA), abscisic acid (ABA), 12-oxo-phytodienoic acid (OPDA), putrescine (Put), and spermine and increases in jasmonic acid (JA), jasmonoyl-isoleucine, trans-cinnamic acid, and salicylic acid contents in shoots at the fourth subculture compared to the first. The ornithine decarboxylase enzyme preferentially functions in the Put biosynthesis pathway, and it was related to the highest AR formation in shoots at the first subculture. Down-accumulation of the auxin-binding protein ABP19a in shoots from the fourth subculture compared to the first subculture was related to a decrease in both the IAA content and AR formation. In addition, down-accumulation of glucose-6-phosphate isomerase; glutamine synthetase leaf isozyme, chloroplastic; 5-methyltetrahydropteroyltriglutamate-homocysteine methyltransferase; l-ascorbate peroxidase, cytosolic; monodehydroascorbate reductase; and 2-Cys peroxiredoxin BAS1-like, chloroplastic and up-accumulation of caffeoyl-CoA O-methyltransferase 1 and isoforms of peroxidase 4 proteins in shoots from the fourth relative to the first subculture were associated with a reduction in AR formation. These results showed that the understanding of hormonal and molecular mechanisms related to the potential of AR formation in shoots under successive subcultures is relevant to improving large-scale plantlet production in C. fissilis.

Melatonin treatment induces chilling tolerance by regulating the contents of polyamine, γ-aminobutyric acid, and proline in cucumber fruit

The mechanism of melatonin (MT) induced chilling tolerance in harvested cucumber fruit was investigated at commercial maturity. In this study, cucumber fruits were treated with 100 µmol L–1 MT at 4°C and 90% relative humidity for 15 d of storage. In comparison with the control, cucumber treatment with MT resulted in reduced chilling injury (CI), decreased electrolyte leakage and enhanced firmness. The fruits treated with MT showed higher chlorophyll contents in storage conditions with suppressed chlorophyllase enzyme activity. MT treatment increased arginine decarboxylase (ADC) and ornithine decarboxylase (ODC) enzyme activities. Moreover, enhanced expression of the Cucumis sativus ADC (CsADC) and C. sativus ODC (CsODC) genes resulted in the accumulation of polyamine contents. Similarly, proline levels exhibited higher levels among treated fruits. Meanwhile, the proline synthesizing enzymes △1-pyrroline-5-carboxylate syntheses (P5CS) and ornithine aminotransferase (OAT) were significantly increased, while a catabolic enzyme of proline dehydrogenase (PDH) activity was inhibited by treatment. In addition, MT induced expression of C. sativus OAT (CsOAT) and C. sativus P5CS (CsP5CS) genes. Cucumber fruits treated with MT also exhibited higher γ-aminobutyric acid (GABA) content by enhanced GABA transaminase (GABA-T) and glutamate decarboxylase (GAD) enzyme activities and a higher C. sativus GAD (CsGAD) gene expression. To sum up, the results show that MT treatment enhanced chilling tolerance, which was associated with the regulation of polyamines, as well as proline and γ-aminobutyric acid.

Optimization of γ-Aminobutyric Acid (GABA) Accumulation in Germinating Adzuki Beans (Vigna angularis) by Vacuum Treatment and Monosodium Glutamate, and the Molecular Mechanisms.

This study aimed to investigate the optimal hypoxic and monosodium glutamate (MSG) stress conditions for the enrichment of γ-Aminobutyric acid (GABA) in germinating adzuki beans and to reveal the potential underlying molecular mechanisms of GABA accumulation. Using single-factor experiments and response surface model, we investigated the effects of germination time, germination temperature, vacuum time, and MSG concentration on GABA contents, and further explored the activity and gene expression of glutamate decarboxylase (GAD) and polyamine oxidase (PAO) critical rate restriction enzymes during GABA synthesis. The optimal soaking temperature, soaking time, and pH conditions were 35°C, 16 h, and 5, respectively. Furthermore, the optimal germination conditions for optimal GABA enrichment were 48 h, 1.99 mg/ml MSG concentration, germination temperature of 31.49°C, and vacuum time of 15.83 h. Under such conditions, the predicted GABA concentration was 443.57 ± 7.18 mg/100 g, with no significant difference between the predicted and experimental data. The vacuum + MSG (FZM) treatment has a maximum contribution rate of GABA to 38.29%, which significantly increase GABA content, and the increase was associated with increased GAD and PAO activity. In addition, MSG in combination with vacuum treatment could significantly induce VaGAD4 and VaGAD6 genes in 2 days germination of adzuki beans. According to the results of the present study, vacuum + MSG treatment is an effective approach to enhancing GABA accumulation in germinating adzuki beans, which could be employed in enhancing the functional quality of germinating adzuki beans.

Cell surface display of Neurospora crassa glutamate decarboxylase on Escherichia coli for extracellular Gamma-aminobutyric acid production from high cell density culture

GABA is produced from the decarboxylation of glutamate by glutamate decarboxylase enzyme (GadB). Various hetrologous GadB enzymes has been indentified and introduced in Escherichia coli to improve the productivity of GABA. In this study, cell surface display strategy was employed by displaying Neurospora crassa glutamate decarboxylase (GadB) on the surface of Escherichia coli using OmpC as an anchoring motif. To construct the display system, the GadB was fused to truncated C-terminus of OmpC and expressed in E. coli. The recombinant E. coli displaying GadB has produced 3.03 g/L of GABA from 10 g/L glutamate within 12 h of culture period. When the GadB display system was cultured at high cell density, 100% GABA yeild was achieved from 50 g/L of glutamate. The results suggest that the GadB expression on the E. coli surface is stable and effective on GABA production. The surface display strategy can be applied on other potential enzyme candidates and used in Industry for bulk chemical production.

Decarboxylation of Aromatic Carboxylic Acids by the Prenylated-FMN-dependent Enzyme Phenazine-1-carboxylic Acid Decarboxylase

Phenazine-1-carboxylic acid decarboxylase (PhdA) is a member of the expanding class of prenylated-FMN-dependent (prFMN) decarboxylase enzymes. These enzymes have attracted interest for their abilit...

Directed evolution of prenylated FMN-dependent Fdc supports efficient in vivo isobutene production

Isobutene is a high value gaseous alkene used as fuel additive and a chemical building block. As an alternative to fossil fuel derived isobutene, we here develop a modified mevalonate pathway for the production of isobutene from glucose in vivo. The final step in the pathway consists of the decarboxylation of 3-methylcrotonic acid, catalysed by an evolved ferulic acid decarboxylase (Fdc) enzyme. Fdc belongs to the prFMN-dependent UbiD enzyme family that catalyses reversible decarboxylation of (hetero)aromatic acids or acrylic acids with extended conjugation. Following a screen of an Fdc library for inherent 3-methylcrotonic acid decarboxylase activity, directed evolution yields variants with up to an 80-fold increase in activity. Crystal structures of the evolved variants reveal that changes in the substrate binding pocket are responsible for increased selectivity. Solution and computational studies suggest that isobutene cycloelimination is rate limiting and strictly dependent on presence of the 3-methyl group.

3-O-Acetyl-11-keto-β-boswellic acid ameliorates chronic unpredictable mild stress induced HPA axis dysregulation in relation with glutamate/GABA aberration in depressive rats.

Overt expression of brain glucocorticoid receptor (GR) leads to elevation of glutamate release causes cerebral excitotoxicity which in turn produce neuropsychological disorders. The aim of our work is to study the consequence of 3-O-Acetyl-11-keto-β-boswellic acid (AKBA) on chronic unpredictable mild stress (CUMS) induced HPA axis dysregulation in relative to glutamate and GABA irregularity in depressive rats. AKBA (5, 10 &15mg/kg) was administered for 28days parallel with CUMS induction in rats. Behavioural studies, tail suspension test (TST), open field exploratory (OFT) and forced swim test (FST) were performed. Biochemical studies including plasma corticosterone, glutamate GABA and glutamic acid decarboxylase (GAD) enzyme activity were studied. Glucocorticoid receptor expression and brain histology were studied to observe the effect of AKBA. CUMS induction results in depressive state of the animals were confirmed by the sucrose preference test. The administration of AKBA significantly reduced the immobility time and improved the exploratory behaviour. Plasma corticosterone and brain glutamate level was decreased and GABA level were increased significantly evident with GAD activation in AKBA-treated animals, further confirmed with decreased GR expression improves architecture of prefrontal cortex region. Correlation study illustrates behavioural improvements undeviating the biochemical alteration and GR expression after AKBA treatment during depression. AKBA significantly reversed the CUMS-induced glutamate/GABA abnormalities through the adaptation of central HPA axis regulation. Hence this study concludes that AKBA can be a better alternative to treat depressive disorders.

Optimization of Growth Conditions for Oxalate Decarboxylase Production from Pseudomonas sp. OXDC12 and in vitro Inhibition of Calcium Oxalate Crystallization by Oxalate Decarboxylase

Background: Kidney stones have become a common disease worldwide, and their incidence and recurrence rates have drastically increased over the past few decades. Oxalate decarboxylase (OxDC) enzyme which catalyzes the disproportionation reaction of oxalate monoanions into formate and CO2 could exhibit significant potential in the treatment of hyperoxaluria. Objective: The present work describes isolation and screening of new OxDC producing bacterial strain from oxalate-rich soils and one-factor-at-a-time (OFAT) and response surface methodology (RSM) statistical approaches were used to optimize the production media to obtain an improved intracellular OxDC production. Methods and Results: An OxDC producing bacterial strain, isolated from spinach soil sample(s), was identified to be Pseudomonas sp. OXDC12 by 16S rRNA sequencing. The OFAT approach was used to determine the effect of supplementation of carbon, nitrogen, and other physical conditions like pH, temperature, etc. on intracellular OxDC production by Pseudomonas sp. OXDC12. The three factors screened by Plackett Burman design (PBD) were further used by the central composite design (CCD) approach of RSM to determine their interactive effects on OxDC production. The anti-urolithiatic activity of the enzyme OxDC was determined by carrying out in vitro calcium oxalate crystallization in the presence and absence of OxDC. The factorial values selected by 23 CCD for OxDC were temperature 30ºC, manganese ion concentration 5 mmol l-1, and inoculum size 3.25% (v/v). The highest predicted value of OxDC was 5.7 U ml-1 while the actual value obtained was 6.7 U ml-1 which was 79.1% and 2.92 fold greater than the initial activity of OxDC produced by Pseudomonas sp. OXDC12. As depicted by the light micrographs, OxDC displayed a significant reduction in the crystallization and formation of calcium oxalate stones as compared to the control under in vitro conditions. Conclusion: OFAT and RSM statistical optimization approaches led to improved OxDC production with a final activity of 6.7 U/mL and a 2.92 fold increase in the enzyme activity. The study suggests that OFAT and RSM optimization approaches significantly enhanced OxDC production from Pseudomonas sp. OXDC12. The enzyme may serve as a potential therapeutic agent for hyperoxaluria or kidney stones as it significantly inhibited the formation of calcium oxalate crystals under in vitro conditions.

Unveiling Complete Ethanol Oxidation through a Hybrid Enzymatic and Organic Catalyst Cascade in an Electrochemical Micro-Reactor Device

Biological fuel cells have been extensively reported as a efficient and sustainable device to convert chemical energy to electrical energy [1]. Biofuel cells (BFCs) are alternative energy sources which use enzymes (enzymatic biofuel cell) or microorganisms (microbial fuel cell) as the electrocatalysts instead of the traditional noble metal catalysts. Ethanol is a fuel that can be used in combined enzymatic fuel cell due to its high energy density (8.03 kW h kg -1), low emission, and storage security [2]. The advantage of enzymes is that they exhibit high specificity and turnover rate, however, mimicking the natural metabolism from ethanol through electrometabolic pathways is enormously difficult. To overcome the limitations of using multiple enzymes to achieve the complete ethanol oxidation, a small molecule organic catalyst can be employed. TEMPO (2,2,6,6-tetramethylpiperidin-1-yl) oxyl) is a small organic catalyst that does not present limited substrate specificity such as enzymes face, due to its ability to oxidize oxygen, nitrogen and sulfur-containing functional groups. Thus, hybrid system enables improvement at the rate of electrocatalytic oxidation of several biofuels [2 - 4]. Herein, we described the development of a hybrid catalytic architecture combining an organic oxidation catalyst, 4-amino-TEMPO (TEMPO-NH2), and a recombinant enzyme, oxalate decarboxylase (OxDc) to catalyze complete ethanol electrooxidation. The catalyst system was coupled with a novel small scale electrolysis cell utilizing polycaprolactone (PCL) and carbon composite electrodes for bulk electrolysis. Electrochemical meansurements and product detection by nuclear magnetic resonance (NMR) after 12 hours of electrolysis have shown for the first time that an organic catalyst and decarboxylase enzyme enhances the overall steps in the cascade operating at acidic pH enabling enhanced electrochemical oxidation of ethanol to CO2. To validate the reaction sequence, a novel micro-reactor device enabled low-cost product analysis and allows for high analyte sensitivity. Overall, this work demonstrated that bi-cascade of TEMPO-NH2 and OxDc can be combined in an electrochemical cell to synergistically catalyze the oxidation of substrates that make up the steps of the ethanol oxidation cascade and is promising for a wide array of applications, including biosensors, environmental monitoring, and biofuel cells.