Enzyme and Microbial Technology最新文献

{"title":"Mechanistic investigation of the enhanced catalytic activity of B. sub lipase A mutant I157V and its application in biodiesel production","authors":"Zhong Ni,&nbsp;Jia Zhao,&nbsp;Huimin Zhou,&nbsp;Huayun Jia,&nbsp;E-Bin Gao,&nbsp;ZhongJian Guo,&nbsp;Zhaoyang Hu,&nbsp;Shangshang Ma,&nbsp;Yong Chen","doi":"10.1016/j.enzmictec.2025.110798","DOIUrl":"10.1016/j.enzmictec.2025.110798","url":null,"abstract":"<div><div>Optimizing lipases for the transesterification of long-chain fatty acid esters plays a vital role in enhancing biodiesel yields, especially from recycled cooking oils. In this study, we applied error-prone PCR to create a diverse mutagenesis library based on <em>Bacillus subtilis</em> lipase A(BSLA). The I157V mutant, selected through 96-well high-throughput screening, exhibited over sixfold greater catalytic efficiency (<em>k</em>_cat/<em>K</em>_m) than the wild-type enzyme. Insights from molecular docking and dynamic simulations pointed to the mutation’s effect on aligning substrates more effectively within the enzyme’s catalytic center. This alignment minimized non-productive binding modes and reinforced the enzyme-substrate interactions, which in turn boosted catalytic output. In biodiesel production using waste cooking oil as feedstock, the I157V mutant achieved a methyl ester yield of 87 % (w/w) after 10 h, representing an ∼45 % increase compared to 60 % (w/w) for the wild type. These findings not only advance enzyme engineering for industrial biocatalysis but also underline the potential of targeted mutations in supporting sustainable energy solutions.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110798"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145682009","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":"PhcK mediates transport of a β-5-type lignin-derived dimer in Sphingobium lignivorans SYK-6","authors":"Mitsuru Kawazoe ,&nbsp;Masaya Fujita ,&nbsp;Shojiro Hishiyama ,&nbsp;Naofumi Kamimura ,&nbsp;Eiji Masai","doi":"10.1016/j.enzmictec.2025.110784","DOIUrl":"10.1016/j.enzmictec.2025.110784","url":null,"abstract":"<div><div>Lignin is a major component of plant cell walls; however, its heterogeneous and complex structure has hindered its efficient utilization. Recently, strategies that combine chemical pretreatment with microbial conversion to produce valuable chemicals have attracted attention. To develop an ideal microbial platform for this purpose, it is essential to elucidate the complete bacterial catabolic system for lignin-derived aromatic compounds. Here, we identified an inner membrane transporter gene involved in the uptake of a metabolite of dehydrodiconiferyl alcohol (DCA), a lignin-derived β-5 dimer, in <em>Sphingobium lignivorans</em> SYK-6. SLG_12820 (<em>phcK</em>) was found to encode a major facilitator superfamily transporter belonging to the endosomal spinster family and is regulated by PhcR, a transcriptional regulator of DCA catabolism genes. Through mutant analysis and a specific uptake assay based on PhcR effector recognition, we demonstrated that PhcK functions as an inner membrane transporter that specifically imports the DCA metabolite, 3-(2-(4-hydroxy-3-methoxyphenyl)-3-(hydroxymethyl)-7-methoxy-2,3-dihydrobenzofuran-5-yl)acrylic acid.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110784"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145622367","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":"A unique highly efficient, thermostable and multi-substrate specific galactanase (AtGH53) from Acetivibrio thermocellus cleaving both β (1,4)- and β (1,6)- linked galactans","authors":"Shreya Biswas ,&nbsp;Ardhendu Mandal ,&nbsp;Carlos M.G.A. Fontes ,&nbsp;Arun Goyal","doi":"10.1016/j.enzmictec.2025.110801","DOIUrl":"10.1016/j.enzmictec.2025.110801","url":null,"abstract":"<div><div>The gene encoding endo-<em>β</em>-1,4-galactanase, <em>At</em>GH53 from <em>Acetivibrio thermocellus</em> was cloned, expressed and the encoded soluble protein was biochemically characterized. Purified <em>At</em>GH53 showed molecular mass of approximately, 36 kDa, an optimum temperature 70°C and half-life of 15 h at 70°C. <em>At</em>GH53 displayed stability in acidic and alkaline pH ranges, with an optimum pH 7.5. <em>At</em>GH53 activity increased over 30 % by Ni or Co²⁺ ions. <em>At</em>GH53 exhibited broad substrate specificity, displaying the highest activity with potato pectic-galactan with <em>V</em>_<em>max</em> of 1432 U.mg⁻¹ and <em>K</em>_<em>M</em> of 1.2 mg.mL⁻¹ . TLC and HPLC analyses of potato galactan hydrolysis by <em>At</em>GH53 showed initially the endo-lytic cleaving property and later shifting to exo-lytic mode. This was confirmed by the release of β-1,4-linked galacto-oligosaccharides of higher degrees of polymerization (DP&gt;3) from potato galactan in first 2 h, followed by accumulation of galactobiose and galactose up to 24 h. In contrast, hydrolysis of larch-arabinogalactan by <em>At</em>GH53 (specific activity, 148.6 U.mg⁻¹) resulted in the release of β-1,6-galactobiose from branches of the polymer. This indicated that <em>At</em>GH53 also exhibits lower efficiency in hydrolyzing β-1,6-galactan of arabinogalactan <em>via</em> an exo-mode of action. The broad substrate specificity, pH stability and thermostability of <em>At</em>GH53 make it a versatile enzyme for biotechnological applications.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110801"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145733640","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":"Structure and functional role of the SLHs-CBM54 tandem in bacterial multimodular glycoside hydrolases","authors":"Oksana V. Berezina ,&nbsp;Alina I. Selimzyanova ,&nbsp;Kirill V. Gordeev ,&nbsp;Wolfgang H. Schwarz ,&nbsp;Natalia A. Lunina","doi":"10.1016/j.enzmictec.2025.110787","DOIUrl":"10.1016/j.enzmictec.2025.110787","url":null,"abstract":"<div><div>Carbohydrate-binding modules (CBMs) enhance enzymatic degradation of polysaccharides by improving substrate binding. CBM54, a family of carbohydrate-binding modules, is found in surface-displayed multimodular glycoside hydrolases (MGHs) of gram-positive bacteria. These modules bind insoluble polysaccharides, including chitosan, chitin, xylan, cellulose, and fungal cell wall β-glucans. In MGHs, the CBM54 module is invariably positioned downstream of three S-layer homology modules (SLHs), which anchor the enzyme to the bacterial cell surface. The SLHs-CBM54 tandem promotes bacterial adherence to substrates, concentrating hydrolytic enzymes at the interface and facilitating efficient uptake of soluble degradation products. CBM54 contains a cleavage site that divides it into two structurally distinct parts, which remain associated via hydrogen bonding. The processing of CBM54 promotes bacterial detachment from the substrate and their migration toward new nutrient sources. Beyond MGHs, the SLHs-CBM54 tandem occurs in many other bacterial proteins with uncharacterized functions. SLH modules may self-assemble into 2D arrays on synthetic supports, enabling their use as matrices for protein nucleation and crystal growth. The SLHs-CBM54 tandem fusion with functional modules offers an innovative approach to surface modification, with broad biomedical and biotechnological applications. Due to its versatile architecture, this system holds promise for antigen display, vaccine development, and enhanced polysaccharide degradation processes.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110787"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145603094","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":"Sensitive and specific fluorometric assay for assessment of glyoxalase II enzymatic activity in microbial samples and biological tissue","authors":"Mohammed Alaa Kadhum,&nbsp;Mahmoud Hussein Hadwan","doi":"10.1016/j.enzmictec.2025.110783","DOIUrl":"10.1016/j.enzmictec.2025.110783","url":null,"abstract":"<div><div>This study presents a novel fluorescence-based assay for quantifying Glyoxalase II (Glo II) enzymatic activity, using N-(9-Acridinyl)maleimide (NAM) as a fluorescent probe. The assay is designed to measure glutathione (GSH) production resulting from the hydrolysis of S-<span>D</span>-lactoylglutathione by Glo II, providing a sensitive and reliable method for assessing enzyme activity across various biological samples. The protocol involves incubating Glo II samples at 37 °C, then adding NAM, which reacts with thiol groups to form a fluorescent adduct. The fluorescence intensity is measured at excitation and emission wavelengths of 360 nm and 432 nm, respectively, allowing for precise quantification of Glo II activity. The NAM-Glo II method demonstrates exceptional sensitivity and specificity, with limits of detection (LOD) and quantification (LOQ) of 0.01 U/L and 0.033 U/L, respectively. This high sensitivity is crucial for accurately measuring Glo II activity in diverse bacterial strains, where enzyme levels may vary. Comparative studies with established methods reveal that the NAM-Glo II assay consistently yields results comparable to, and in some cases superior to, those obtained using UV-based techniques. Notably, the method effectively minimizes interference from common biomolecules, such as amino acids and carbohydrates, which can confound traditional assays. The NAM-Glo II method is a reliable, sensitive tool for quantifying Glo II activity, crucial for neurological and microbial studies. It enables accurate enzyme measurement, reveals higher activity in <em>E. coli</em>, aids bacterial metabolism research, and supports insights into detoxification, resistance, and targeted antimicrobial therapies.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110783"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145578556","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-enzyme co-fermentation of low-grade tobacco: Metagenomics and metabolomic insights into flavor formation","authors":"Ming Shu ,&nbsp;Huijie Xue ,&nbsp;Yang Yang ,&nbsp;Xiao Zhang ,&nbsp;Shitou Li ,&nbsp;Tengfei Bian ,&nbsp;Kailong Yuan ,&nbsp;Chunping Xu","doi":"10.1016/j.enzmictec.2025.110803","DOIUrl":"10.1016/j.enzmictec.2025.110803","url":null,"abstract":"<div><div>Microbial-enzyme co-fermentation effectively enhances the quality of low-grade tobacco leaves quality, but the underlying mechanisms of flavor formation remain unclear. This study investigated the dynamics and relationships of microbial communities and volatile aroma metabolites during low-grade tobacco leaves fermentation through metagenomics and headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME-GC-MS). Results showed that during microbial-enzyme co-fermentation, the tobacco leaves fermented for four days (D4) exhibited the highest levels of total sugars and reducing sugars, the peak total content of aroma metabolites, and the best sensory quality. <em>Pseudomonadota</em>, <em>Bacillota</em>, and <em>Ascomycota</em> were dominant microorganisms during fermentation. During the initial stage (D1–D4), <em>Saccharomyces</em> was the dominant genus, which was subsequently displaced by <em>Pantoea</em> at D5. This microbial succession coincided with a decline in sensory quality, indicating its crucial role in shaping flavor evolution during co-fermentation. During microbial-enzyme co-fermentation process, a total of 46 volatile metabolites were detected in low-grade tobacco leaves. Among them, seven esters with high variable important in projection values and strong microbial correlations were identified as characteristic aroma metabolites, including ethyl phenylacetate, benzyl acetate, phenylethyl acetate, ethyl myristate, ethyl palmitate, ethyl oleate, and methyl linolenate. Gene function annotation revealed carbohydrate metabolism was the most abundant, followed by amino acid metabolism. Spearman correlation analysis elucidated the formation mechanism of characteristic ester metabolites. Specifically, short-chain esters correlated with glycerolipid and amino acid metabolism, while long-chain esters linked to glycolysis and fatty-acid biosynthetic pathways.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110803"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145733639","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 and optimization of an engineered E. coli platform for nitrotryptophan biosynthesis","authors":"Haiying Lin ,&nbsp;Man Zhang ,&nbsp;Wenbin Kang ,&nbsp;Jianru Pan","doi":"10.1016/j.enzmictec.2025.110785","DOIUrl":"10.1016/j.enzmictec.2025.110785","url":null,"abstract":"<div><div>Nitrotryptophan and its derivatives are valuable building blocks for synthesizing bioactive compounds and functional materials. This study reports the development of an efficient and novel bio-catalytic bioreactor in <em>Escherichia coli</em> capable of the direct aromatic nitration of tryptophan, enabling the synthesis of nitrotryptophan isomers. The biosynthetic pathway incorporates a self-sufficient P450 enzyme (TB14, consisting of TxtE-linker14-BM3R) from Streptomyces for the direct insertion of a nitro group into the indole ring of <span>L</span>-tryptophan. This process is supported by a nitric oxide synthetase (BsNOS) from <em>Bacillus subtilis</em> or its chemical alternative, sodium nitroprusside (SNP), to produce nitric oxide (NO) from <span>L</span>-arginine, which facilitates the direct nitration. As both TB14 and BsNOS require the reductant NADPH for their respective biochemical reactions, a glucose dehydrogenase (GDH) from Bacillus subtilis was included in the experimental design to ensure NADPH regeneration within the system.The initial engineered strain produced 133.2 mg/L of nitrotryptophan in TB medium. Through systematic optimization, including pathway balancing, fermentation condition enhancement, and elimination of competing metabolic pathways, the final titer was successfully increased to 209.9 mg/L within 48 h. This work establishes a robust platform for the microbial production of valuable nitroaromatic compounds and provides key insights for future biocatalytic nitration strategies.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110785"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145578555","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":"Recombinant expression and nucleotide hydrolysis activity of NTPDase 4 from Trichomonas vaginalis","authors":"Magno Sinval Pereira Ribeiro ,&nbsp;Laura Cipolatto da Rosa ,&nbsp;Giulia Bongiorni Galego ,&nbsp;Mateus Dias de Oliveira ,&nbsp;Carolina Wicteky Totti ,&nbsp;Rogerio Margis ,&nbsp;Geancarlo Zanatta ,&nbsp;Tiana Tasca","doi":"10.1016/j.enzmictec.2025.110786","DOIUrl":"10.1016/j.enzmictec.2025.110786","url":null,"abstract":"<div><div><em>Trichomonas vaginalis</em> is the etiologic agent of trichomoniasis, the non-viral sexually transmitted infection most prevalent in world. It is important to investigate biochemical aspects of the parasite that contribute to our understanding of the biology and applications in the treatment and diagnosis of the infection. The nucleoside triphosphate diphosphohydrolase (NTPDase) is an enzyme that hydrolyses extracellular adenine and guanine nucleotides, forming nucleosides adenosine and guanosine. This is important for parasite survival through the purine salvage pathway, since adenosine is the precursor for the entire purine nucleotides pool in <em>T. vaginalis</em>. Herein we expressed TvNTPDase4 in the bacterial system <em>Escherichia coli</em>. Our data demonstrate that the enzyme is active, being able to hydrolyze ATP, ADP and AMP at a concentration of 10 μg of purified protein/reaction. The inhibitors gadolinium and adenosine 5′-[α,β-methylene]diphosphate (AMPCP) inhibited the hydrolysis of rTvNTPDase4. The inhibition of ATPase/ADPase activity was more effective with gadolinium, while the inhibition of AMPase activity was more effective with AMPCP. The enzyme rTvNTPDase4 was not cytotoxic to HMVII cells. In molecular dynamics, we observed that the ability of TvNTPDase4 to hydrolyze ATP, ADP, and AMP substrates occurs through direct interactions with the apyrase-conserved regions (ACR), especially ACR1 and ACR4. In this work, we did not find any candidate sequence for ecto-5′-nucleotidase (E-5′-N) in <em>T. vaginalis</em>, which leads us to believe that the parasite does not have this enzyme in its proteomic repertoire. Finally, we report that rTvNTPDase4 expressed and purified from a bacterial is active and has potential for biotechnological applications.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110786"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145578557","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":"Peach gum-derived immobilization of Klebsiella sp. cells combined with nanoparticle addition for robust enhancement of biohydrogen production","authors":"Qin Zhang ,&nbsp;Qianzi Cheng ,&nbsp;Juanjuan Cao ,&nbsp;Cancan Zhang ,&nbsp;Bo Wu ,&nbsp;Yanbin Li ,&nbsp;Sejabodile Precious Moanokeng","doi":"10.1016/j.enzmictec.2025.110800","DOIUrl":"10.1016/j.enzmictec.2025.110800","url":null,"abstract":"<div><div>This study developed a peach-gum-derived immobilization of <em>Klebsiella</em> sp. cells combined with nanoparticles (NPs) to enhance biohydrogen production. The optimal ratio of peach gum (PG) to sodium alginate was determined to be 1.5:1 for effective immobilization of <em>Klebsiella</em> sp. cells, which was further improved by the addition of magnetite NPs and NiFe₂O₄ NPs (NFNPs). The results showed that the Im + NFNP 20 mg/L treatment was optimal, achieving the highest cumulative hydrogen production of 5996.35 ± 0.29 mL/L at 120 h, which was 1.89 times higher than the control treatment. Notably, this treatment resulted in the maximum <em>Y</em>_{<em>H</em><em>2/S</em>} (128.42 ± 2.05 mL/g) and the highest peak number of viable cells ((2.91 ± 0.03) * 10⁹ cfu/mL), as well as enhanced peak activities of hydrogenase and formate-hydrogen lyase. Furthermore, the combination of immobilization and optimum NFNP addition noticeably promoted the expressions of several hydrogenase and formate-hydrogen lyase related genes and altered the distribution of soluble metabolites, which enhanced the hydrogenase- and formate-hydrogen lyase-catalyzed pathways. Thus, we propose a mechanism in which PG-derived immobilization facilitates the entry of NPs into <em>Klebsiella</em> sp. cells, and elucidate the molecular basis for the efficient cellular uptake and distribution of NFNPs in <em>Klebsiella</em> sp. cells.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"194 ","pages":"Article 110800"},"PeriodicalIF":3.7,"publicationDate":"2026-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145713783","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":"Modulation of ADH activity in Zymomonas mobilis provides significant flux redirection away from ethanol","authors":"Arun Thapa,&nbsp;Ngangom Pravina Devi,&nbsp;Ashish Misra","doi":"10.1016/j.enzmictec.2025.110777","DOIUrl":"10.1016/j.enzmictec.2025.110777","url":null,"abstract":"<div><div><em>Zymomonas mobilis</em> has an extremely high specific glucose uptake rate and produces ethanol at high yields and productivities. For synthesizing products beyond ethanol in <em>Z. mobilis,</em> reducing ethanol yields is a major challenge. Previous efforts have sought to decrease PDC activity to minimize ethanol formation. Here, we sought to modulate ADH (encoded by <em>adhA</em> and <em>adhB</em> genes) activity to redirect flux away from ethanol. We found that deletion of <em>adhB</em> combined with co-expression of a heterologous NAD(P) regenerating enzyme (<span>L</span>-LDH) diverted more than 50 % of carbon flux away from ethanol formation. The sequential deletion of <em>adhA</em> in the <em>adhB</em>-deleted strain led to ∼90 % reduction in ADH activity. During batch growth, the strain showed ∼90 % reduction in ethanol titres (27 mM) compared to WT (209 mM) and significant flux redirection toward <span>L</span>-LA formation (169 mM). The results demonstrate that modulation of ADH activity by deletion of <em>adhA</em> and <em>adhB</em> genes is an effective strategy for rediverting flux toward products beyond ethanol in <em>Z. mobilis.</em></div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"193 ","pages":"Article 110777"},"PeriodicalIF":3.7,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145548408","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}