Enzyme and Microbial Technology最新文献

{"title":"Identification of a 3-deoxy-D-manno-octulosonic acid kinase of lipid A in Vibrio parahaemolyticus","authors":"Lingyan Chen ,&nbsp;Danyang Huang ,&nbsp;Xinrui Zhang ,&nbsp;Hongchen Yin ,&nbsp;Xiaoyuan Wang","doi":"10.1016/j.enzmictec.2025.110775","DOIUrl":"10.1016/j.enzmictec.2025.110775","url":null,"abstract":"<div><div><em>Vibrio parahaemolyticus</em> is a major seafood-associated foodborne pathogen whose lipopolysaccharide (LPS) plays an important role in virulence and antimicrobial resistance. The LPS of <em>V. parahaemolyticus</em> contains a single 3-deoxy-D-manno-octulosonic acid (Kdo) sugar with phosphorylation. Previously, we have characterized the gene <em>VP_RS01035</em> is responsible for the addition of Kdo; in this study, we characterized another gene <em>VP_RS00960</em> which is responsible for the Kdo phosphorylation of <em>V. parahaemolyticus</em> LPS. To investigate its function, we first constructed an LPS-deficient <em>Escherichia coli</em> WH600 strain using CRISPR/Cas9. Heterologous expression of <em>VP_RS01035</em> alone or in combination with <em>VP_RS00960</em> yielded recombinant strains WH600/pB1–1 and WH600/pB2–12, respectively. Analysis of total lipids from the recombinant strains by thin-layer chromatography and high-performance liquid chromatography-tandem mass spectrometry demonstrated that the <em>VP_RS00960</em> gene encodes a Kdo kinase responsible for phosphorylating the 4-OH site of the Kdo sugar in <em>V. parahaemolyticus</em>. To further validate its role, we deleted <em>VP_RS00960</em> gene in <em>V. parahaemolyticus,</em> resulting in mutant Δ<em>RS00960</em>. Notably, Δ<em>RS00960</em> failed to produce polysaccharide-linked lipid A, although free lipid A synthesis remained unaffected. Furthermore, defective long-chain LPS assembly compromised outer membrane integrity, increasing permeability and hydrophobicity while reducing biofilm formation. Consequently, Δ<em>RS00960</em> exhibited heightened susceptibility to membrane-targeting antibiotics, such as erythromycin and novobiocin. Macrophage infection assays using RAW264.7 cells revealed that <em>VP_RS00960</em> deletion attenuated bacterial pathogenicity. These findings enhance the understanding of the pathogenicity and drug resistance of <em>V. parahaemolyticus</em>, and provide novel insights and strategies for addressing antibiotic resistance and food safety challenges posed by <em>V. parahaemolyticus</em>.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110775"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145463985","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Highly efficient spermidine production system in Escherichia coli BL21 (DE3) based on precursor metabolic modules optimization and carboxyaminopropylagmatine pathway construction","authors":"Jie Wang ,&nbsp;Linbo Gou ,&nbsp;Di Liu ,&nbsp;Shengfang Wu ,&nbsp;Xiuwen Zhou ,&nbsp;Tai-Ping Fan ,&nbsp;Long Wang ,&nbsp;Yujie Cai","doi":"10.1016/j.enzmictec.2025.110763","DOIUrl":"10.1016/j.enzmictec.2025.110763","url":null,"abstract":"<div><div>Spermidine, a naturally occurring polyamine compound, has garnered significant attention due to its versatile physiological functions, including induction of cellular autophagy, antioxidant activity, and maintenance of mitochondrial homeostasis. In this study, we established a novel spermidine biosynthesis system in <em>E. coli</em> BL21 (DE3) by heterologously introducing the carboxyaminopropylagmatine (CAPA) pathway derived from cyanobacterium. To enhance precursor supply, we overexpressed key enzymes in the L-aspartate β-semialdehyde and agmatine branch metabolic pathways within the precursor metabolic module, while simultaneously knocking out competing metabolic pathways to redirect metabolic flux toward spermidine biosynthesis. To address the challenge of intracellular spermidine accumulation and its associated cytotoxicity, the high-efficiency spermidine efflux pump protein AmvA from <em>Acinetobacter baumannii</em> was heterologously expressed in <em>E. coli</em> BL21 (DE3). This engineering strategy enabled efficient extrusion of spermidine from the cells, alleviating the toxic effects of high intracellular spermidine concentrations on the host strain. Through these metabolic and efflux pump engineering modifications, the engineered strain SPD06-P5-P6 was constructed. Following 48 h of shake flask fermentation, SPD06-P5-P6 achieved a spermidine titer of 163.11 mg/L, which further increased to 1164.22 mg/L after 96 h of scale-up cultivation in a 5 L bioreactor.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110763"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145361483","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Development of high-efficiency hybrid strains for cellulolytic enzyme production via interspecific and intergeneric protoplast fusion of Trichoderma and Penicillium species","authors":"Mohammad J. Alsarraf ,&nbsp;Aisha S.M. Al-Wahaibi ,&nbsp;Steven L. Stephenson ,&nbsp;Najla A. Alshaikh ,&nbsp;Fuad Ameen","doi":"10.1016/j.enzmictec.2025.110764","DOIUrl":"10.1016/j.enzmictec.2025.110764","url":null,"abstract":"<div><div>Forty-five fungal strains from decomposing wood, representing eight genera, were isolated. Among them, 27 cellulolytic strains were identified. The genera <em>Trichoderma</em>, <em>Penicillium</em>, and <em>Phanerochaete</em> (6, 2, and 4 isolates, respectively) demonstrated the highest cellulase production. The isolates TW-25, TW-28, and TW-33 exhibited superior enzyme activities, CMCase (5.4–6.5 U/mL), FPase (3.2–3.8 U/mL), pNPCase (2.6–3.0 U/mL), and pNPGase (3.5–4.0 U/mL) and were selected for further study. Internal transcribed spacer (ITS) region sequencing, combined with phenotypic characteristics, identified these strains as <em>Trichoderma</em> sp. TW-25, <em>Trichoderma</em> sp. TW-28, and <em>Penicillium</em> sp. TW-33. Maximum protoplast release was observed in <em>Trichoderma</em> sp. TW-25 (3.6 × 10⁶ protoplasts/mL), followed by <em>Trichoderma</em> sp. TW-28 (3.0 × 10⁶ protoplasts/mL) and <em>Penicillium</em> sp. TW-33 (2.8 × 10⁶ protoplasts/mL). Fusion frequencies were 2.8 × 10⁻³ for TW-25 × TW-28, 2.0 × 10⁻³ for TW-25 × TW-33, and 1.8 × 10⁻³ for TW-28 × TW-33. A total of 13 colonies obtained from TW-25 × TW-28, and 18 from intergeneric fusions (10 from TW-25 × TW-33 and 8 from TW-28 × TW-33). The cellulase activity of the fusants TWF1/1, TWF1/6, and TWF2/5 was the same as TW-25 and the fusants TWF1/3 and TWF3/2 the same as TW-28 while none of the fusants had the cellulase activity of TW-33. Fusants differed from their parental strains in their DNA content (3.25–3.65 µg/mg dry weight) and showed high cellulase activities in general. Among them, TWF1/10 demonstrated the highest enzymatic activity, producing CMCase, FPase, pNPCase, and pNPGase (10.5, 6.5, 5.8, and 7.5 U/mL), respectively, followed by TWF1/13, TWF2/8, TWF2/10, and TWF3/8. DNA banding patterns of TWF1/10, TWF1/13, TWF2/8, TWF2/10, and TWF3/8, analyzed using four RAPD and three ISSR primers, differed from their parental strains, except for ISSR-3 with fusants TWF1/10 and TWF1/13. These variations underscore the effectiveness of interspecific and intergeneric protoplast fusion. The supernatant of the hybrid strain TWF1/10 was concentrated and purified via ultrafiltration, and SDS-PAGE and zymogram assays confirmed its cellulase activity using CMC as the substrate.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110764"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145376592","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Silica immobilized PETase for microplastic bioremediation: Influence of linker peptides on activity","authors":"Sonia Su,&nbsp;Amol D. Pagar,&nbsp;Julie M. Goddard","doi":"10.1016/j.enzmictec.2025.110782","DOIUrl":"10.1016/j.enzmictec.2025.110782","url":null,"abstract":"<div><div>Environmental microplastic leads to bioaccumulation in humans, animals, and plants with potential toxicity. Polyethylene terephthalate (PET) degrading enzymes (PETases) present an opportunity to depolymerize PET in key intervention points, such as wastewater treatment. While PETase has been extensively studied since its discovery and modified for enhanced performance (especially thermostability), knowledge on immobilization for reusability remains limited. This study investigated the effect of linker peptides between functional, active, stable, and tolerant (FAST) PETase and a silica binding protein for immobilization onto silica and the reusability of the enzyme. Linker peptides and a silica binding protein were assembled onto FAST-PETase and expressed in <em>Escherichia coli</em>. The activity of the constructs was tested on PET before and after binding to silica. In the free system, repeating (GGGGS)₃ flexible linker achieved the same activity as FAST-PETase parent enzyme after 48 h of degradation. Once immobilized to silica, repeating (GGGGS)₃ flexible linker preserved 50 % of enzymatic activity, compared to free FAST-PETase, and 80 % compared to its free form. Silica-immobilized enzyme constructs all retain at least 15 % of relative activity compared to the first cycle of use after 5 reuse cycles. Integration of linker peptides between the enzyme and the silica binding peptide had a significant effect on the overall catalytic activity of FAST-PETase and advances our understanding of immobilized PETase for potential recovery and reuse in applications such as wastewater treatment.</div></div><div><h3>Synopsis</h3><div>Minimal research exists on the immobilization of polyethylene terephthalate degrading enzymes for reuse in environmental systems. This study reports the ability of silica immobilized enzyme, with aid of linker peptides, to minimize microplastic contamination from wastewater treatment plants.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110782"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145556535","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Study on bio-imprinted Aspergillus niger lipase cross-linked aggregates and catalytic synthesis of Vitamin E succinate","authors":"Junqing Qian,&nbsp;Zhengze Yang,&nbsp;Aomei Huang,&nbsp;Qian Li,&nbsp;Hui Guo","doi":"10.1016/j.enzmictec.2025.110766","DOIUrl":"10.1016/j.enzmictec.2025.110766","url":null,"abstract":"<div><div>To enhance the esterification and stability of lipase in organic solvents, the bio-imprinted <em>Aspergillus niger</em> lipase combined with cross-linked aggregate immobilization was investigated. The bio-imprinted lipase cross-linked aggregates were applied to the catalytic esterification for the synthesis of Vitamin E succinate in <em>N, N</em>-dimethylformamide (DMF) solution. Lauric acid, serving as a succinic acid analogue, was selected as the bio-imprinting molecule, 0.10 g lauric acid was added to 36 mL of 2.10 mg/mL lipase solution, imprinting 40 mins at pH 8.0, the immobilization yield achieved 91.5 % with cross-linked aggregates by glutaraldehyde. The catalytic activity of the bio-imprinted lipase cross-linked aggregates was significantly enhanced, achieving an esterification yield of 87.4 ± 0.43 % for Vitamin E succinate. Moreover, the bio-imprinted lipase cross-linked aggregates maintained their catalytic activity over five consecutive reaction cycles in DMF. Fluorescence spectroscopy analysis revealed that bio-imprinting promoted the exposure of the lipase active sites, which corresponded with the observed increase in esterification activity. In addition, the mechanism of the substrate analogue-imprinted lipase was characterized. This study provides a theoretical foundation for improving the catalytic esterification performance of lipase as well as a process basis for the enzymatic synthesis of Vitamin E succinate.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110766"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145387388","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mutational study-based identification of high activity O-methyltransferase for the regioselective methylation of epigallocatechin gallate","authors":"Gyeongguk Park ,&nbsp;Byung-Gee Kim ,&nbsp;Joo-Hyun Seo","doi":"10.1016/j.enzmictec.2025.110779","DOIUrl":"10.1016/j.enzmictec.2025.110779","url":null,"abstract":"<div><div>The site-specific methylation of (-)-epigallocatechin-3-<em>O</em>-gallate (EGCG), especially 3`` hydroxyl group in <span>D</span>-ring, significantly enhances its stability and bioavailability in human body. Therefore, the methylation is the way to make EGCG more orally active and potent nutraceutical agents. In this study, for the effective synthesis of the methylated EGCG, <em>S</em>-adenosyl-<span>L</span>-methionine dependent <em>O</em>-methyltransferase (OMT) was studied to synthesize the methylated EGCG. OMT from <em>Bacillus licheniformis</em> and <em>B. megaterium</em> were known to have methylation activity for EGCG. Because OMT from <em>B. licheniformis</em> (Bl_OMT) showed higher activity and regioselectivity between two OMTs, rational design was tried using Bl_OMT. F163W mutant of Bl_OMT showed 2-fold increased initial velocity for the methylation of EGCG than wild type Bl_OMT. This engineering result was further utilized as a basis for the identification of highly active OMT from sequence database. 100 homolog sequences of OMT of <em>B. licheniformis</em> and 100 homolog sequences of OMT of <em>B. megaterium</em>, were collected using BLAST. Multiple alignment of 202 sequences was used to generate subgroups. Four representative sequences from each subgroup were further studied. As a result, OMTs from <em>Thermolongibacillus altinseunsis</em> and <em>B. subtilis</em>, which were from homolog group of Bl_OMT, showed higher activity than Bl_OMT while showing the same high regioselectivity. OMTs from <em>T. altinseunsis</em> and <em>B. subtilis</em> showed <em>k</em>_cat/<em>K</em>_M of 17.4 M⁻¹s⁻¹ and 11.3 M⁻¹s⁻¹, respectively, while Bl_OMT showed 8.7 M⁻¹s⁻¹. Therefore, we could find that phenylalanine residue of the active site of OMT is very important to make strong binding of hydrophobic moiety of substrate and mutation to tryptophan is able to give higher binding strength.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110779"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145548344","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Microbial-catalyzed biotransformation of Soyasapogenol B by Streptomyces griseus ATCC 13273, and Penicillium griseofulvum CICC 40293","authors":"Richa Raj ,&nbsp;Pingping Shen ,&nbsp;Xuewa Jiang ,&nbsp;Jingling Zhang ,&nbsp;Weihang Yao ,&nbsp;Wei Wang ,&nbsp;Jian Zhang","doi":"10.1016/j.enzmictec.2025.110765","DOIUrl":"10.1016/j.enzmictec.2025.110765","url":null,"abstract":"<div><div>This study aims to diversify the structure of Soyasapogenol B (1) and generate novel metabolites through microbial-catalyzed biotransformation by two highly efficient microbial strains, <em>Streptomyces griseus</em> ATCC 13273 and <em>Penicillium griseofulvum</em> CICC 40293. Consequently, ten (2−11) unique bioactive metabolites were isolated. Their structures were determined using 1D/2D NMR and HR-ESI-MS data, revealing multiple tailoring reactions, including oxidation, C-C double bond rearrangement, hydroxylation, and dehydrogenation. This highlights the enzymatic ability of these strains to catalyze specific and diverse regioselective modifications on the Soyasapogenol B scaffold. Therefore, this study demonstrates that microbial-catalyzed biotransformation offers a promising approach to increase the chemical diversity of Soyasapogenol B (1), providing a sustainable alternative to chemical synthesis.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110765"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145361482","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Ovomucoid hydrolysates prepared using alcalase suppress adipogenesis at an early stage of 3T3-L1 cell differentiation","authors":"Seung-Woo Yu ,&nbsp;Hyeong-Jin Kim ,&nbsp;Su-Yeon Song ,&nbsp;Kee-Tae Kim ,&nbsp;Dong Uk Ahn ,&nbsp;Na-Kyoung Lee ,&nbsp;Hyun-Dong Paik","doi":"10.1016/j.enzmictec.2025.110781","DOIUrl":"10.1016/j.enzmictec.2025.110781","url":null,"abstract":"<div><div>The effects of ovomucoid (OVM), a by-product of egg white, and its hydrolysates on adipocyte differentiation and lipid accumulation were investigated. The OVM hydrolyzed using Alcalase® and pepsin was named AH and PH, respectively. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis analysis revealed significant changes in molecular weight of both hydrolysates, with AH showing a higher degree of hydrolysis. AH exhibited a more pronounced inhibitory effect on fat accumulation than PH. In <em>in vitro</em> experiments, AH and PH suppressed lipid accumulation during 3T3-L1 adipocyte differentiation, with AH inhibiting lipid accumulation most effectively. Oil red O staining and triglyceride measurements revealed lipid reduction in AH-treated cells, indicating that AH plays a major role in preventing lipid accumulation in adipocytes. In addition, AH inhibited the expression of lipid transcription factors (CCAAT/enhancer-binding protein alpha (C/EBP-α), peroxisome proliferator-activated receptor gamma (PPAR-γ), and sterol regulatory element-binding proteins (SREBP-1c)), adipogenesis-related factors (fatty acid synthase (FAS) and ACC1), insulin-related factors (insulin receptor substrate (IRS2) and protein kinase B (AKT2)), and lipolysis-related factors (glycerol-3-phosphate acyltransferase (GPAT), CD36, and lipoprotein lipase (LPL)) in a concentration-dependent manner. Specifically, the effect of AH was most pronounced in the early stages of adipocyte differentiation, where it activated AMPK early to associate energy homeostasis and downregulate genes important for cell cycle and lipid formation. This study suggests that OVM hydrolysates prepared using Alcalase® may contribute to the development of new strategies for the obesity treatment market.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110781"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145563223","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Bioconversion of phenolic acids from hydrolyzed sugarcane bagasse into vinyl derivatives by recombinant phenolic acid decarboxylase","authors":"Maitê Bernardo Correia dos Santos ,&nbsp;Vitória Gonçalves Navarrete ,&nbsp;Mohammed Anas Zaiter ,&nbsp;Erike Jhonatan Pereira ,&nbsp;Jorge Enrique Hernández González ,&nbsp;Natália Ellen Castilho de Almeida ,&nbsp;Roberto da Silva ,&nbsp;Maurício Boscolo ,&nbsp;Sérgio Goméz Alonso ,&nbsp;Eleni Gomes","doi":"10.1016/j.enzmictec.2025.110780","DOIUrl":"10.1016/j.enzmictec.2025.110780","url":null,"abstract":"<div><div>Several commercially important vinyl phenolic compounds can be produced by enzymatic decarboxylation of phenolic acids such as ferulic and <em>p</em>-coumaric acids which can be extracted from agro-industrial waste. Phenolic acid decarboxylase is an enzyme that acts in the decarboxylation of these acids to 4-vinylguaiacol and 4-vinylphenol, respectively. In this study, the gene encoding phenolic acid decarboxylase from <em>Klebsiella pneumoniae</em> TD 4.7 was isolated and identified as a 504 bp fragment, encoding a polypeptide of 167 amino acid residues. A 98 % predicted amino acid sequence identity between ferulic acid decarboxylase from other bacteria of the same genus was determined. The gene was successfully expressed in <em>Escherichia coli</em> BL21 (DE3), and the recombinant enzyme was purified as active in absence of cofactor. The protein had a mass of 22-kDa protein, with greater activity at pH 5.5 and 40 °C. The decarboxylase activity was inhibited by Hg^2 + , Zn²⁺, Cu²⁺, and Cd²⁺ ions and increased by 20 % in the presence of Co²⁺. The K_m and V_max values for the recombinant enzyme were estimated at 2.95 mM and 102.10 µmol min⁻¹ mg⁻¹, respectively. The enzyme’s structure was modelled using the structural prediction programs AlphaFold Multimer and SWISS-MODEL, with an RMSD of just 0.7 Å, demonstrating the absence of cysteine and disulfide bonds in the homodimer, with the presence of a high number of lysine residues. The amino acids involved in the catalytic site were Tyr27, Glu134, and Asn23. The Enzyme activity on substrates ferulic and <em>p</em>-coumaric acids extracted from sugarcane bagasse, resulted in 4-vinylguaiacol and 4-vinylphenol, respectively, with conversion yields of 43 % for ferulic acid and 55 % for <em>p</em>-coumaric acid. These data are important in terms of obtaining an enzyme that decarboxylates ferulic and <em>p</em>-coumaric acids obtained from sugarcane bagasse hydrolyzed with similar efficiency, in a single step and without the need for a cofactor, making it an excellent option for bioprocesses using lignocellulosic biomass derivatives.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110780"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145569257","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Overexpression, biochemical characterization, and structural modeling of polyhydroxybutyrate depolymerase from Nocardiopsis dassonvillei","authors":"H. Anjulal ,&nbsp;Aritri Saha ,&nbsp;Vitthal T. Barvkar ,&nbsp;Kshitija Pawar ,&nbsp;Manali Joshi ,&nbsp;Smita S. Zinjarde","doi":"10.1016/j.enzmictec.2025.110767","DOIUrl":"10.1016/j.enzmictec.2025.110767","url":null,"abstract":"<div><div>The ability of <em>Nocardiopsis dassonvillei</em> NCIM 5124 to synthesize polyhydroxybutyrate depolymerase (PHBD) was recently reported. In this investigation, <em>in vitro</em> codon optimized gene synthesis, overexpression, and biochemical characterization of this enzyme along with molecular docking studies are presented. The sequence of the PHBD was inserted in pET-28a(+) along with the PelB_Signal and His₆ tag to generate the recombinant vector pET-Nd-pelB_PHBD. The transformed <em>Escherichia coli</em> BL21(DE3) could produce active PHBD. This enzyme was purified using Ni-NTA affinity chromatography, producing a product with a molecular weight of roughly 50 kDa. The optimum temperature and pH of the recombinant enzyme were 35°C and 8.0, respectively. Triton X100 and Tween 20 inhibited the enzyme activity by 90 %, indicating the role of hydrophobic residues in the active site of the enzyme, as also noted during docking studies. On the basis of the Michaelis-Menten equation, apparent K_m and V_max of recombinant PHBD were found to be 1.782 mg/mL and 4.79 U/mL/min, respectively. Molecular docking studies indicated that the hydrophobic amino acids Cys 39, Ala 40, Cys 77, Phe 158, Met 161, Val 201, Ala 272, and Tyr 273 present in the catalytic site were providing the necessary hydrophobic environment for binding of the ligand. The purified enzyme could also degrade films of PHB and poly (3-hydroxybutyrate-co-3-hydroxyvalerate-co-3-hydroxyhexanoate (PHBVH). As far as we are aware, this is the first report on the overexpression of PHBD from <em>Nocardiopsis</em> sp.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110767"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145387386","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}