Metabolic Engineering Communications最新文献

{"title":"Fine modulation of carbon flow in central carbon metabolism via ribosome-binding site modification in Escherichia coli","authors":"Shogo Sawada,&nbsp;Tatsumi Imada,&nbsp;Hikaru Nagai,&nbsp;Philip Mundt,&nbsp;Fumio Matsuda,&nbsp;Hiroshi Shimizu,&nbsp;Yoshihiro Toya","doi":"10.1016/j.mec.2026.e00270","DOIUrl":"10.1016/j.mec.2026.e00270","url":null,"abstract":"<div><div>Optimization of flux distribution in central carbon metabolism is important to improve the microbial productivity. As the number of precursors required for synthesis differs for each target compound, optimal flux distribution also varies. A library of mutant strains with diverse flux distributions can aid in optimal strain screening. Therefore, in this study, we aimed to construct a library of <em>Escherichia coli</em> strains with stepwise changes in flux distribution by introducing mutations into the ribosome-binding sites of key enzyme genes on its chromosome. We focused on the flux ratios at the glucose-6-phosphate and acetyl-CoA branch points to enhance mevalonate production. Mutations were introduced into the ribosome-binding sites of <em>pgi</em> and <em>gltA</em> to vary the flux ratios of the two pathway branches. Furthermore, a combinatorial repression library comprising 16 strains was constructed by varying <em>pgi</em> and <em>gltA</em> expression at four levels, and a plasmid containing mevalonate synthesis genes was introduced into each strain. Batch cultures were performed to obtain strains with mevalonate titers and yields 2.4- and 3.4-fold higher than those of the parent strain. Overall, our combinatorial suppression library of <em>pgi</em> and <em>gltA</em> facilitated the effective identification of mutants with optimal metabolism for mevalonate production.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"22 ","pages":"Article e00270"},"PeriodicalIF":4.1,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146034884","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Utilization of phosphate-responsive promoters for auto-inducible management of biotechnological processes based on cyanobacteria","authors":"Marvin Amadeus Itzenhäuser ,&nbsp;Franziska Hufnagel ,&nbsp;Carla Klemm ,&nbsp;Kevin Otec ,&nbsp;Minmin Pan ,&nbsp;Stephan Klähn","doi":"10.1016/j.mec.2026.e00278","DOIUrl":"10.1016/j.mec.2026.e00278","url":null,"abstract":"<div><div>Controllable gene expression is essential in microbial biotechnology, yet most systems rely on costly external inducers that limit large-scale applicability. Here, we employed phosphate-responsive promoters controlled by the SphS–SphR phosphate-sensing two-component system as auto-inducible expression systems in cyanobacteria. Specifically, we characterized the promoters of <em>phoA</em>, <em>sphX</em>, and <em>pstS2,</em> which are activated under low phosphate availability and achieved induction folds of up to 13, as well as P<em>urtA</em> and a synthetic promoter P<em>P</em>_<em>i</em><em>-neg</em>, which are repressed under these conditions. Replacement of the ribosome binding site in the native promoter systems further expanded the accessible range of expression levels, with context-dependent increases or decreases depending on the promoter. By systematically adjusting the phosphate concentration and cell density, the timing of gene expression could be precisely controlled. Notably, intracellular phosphate storage during early growth enables transient buffering of external depletion, allowing promoter activation prior to growth limitation. This, in turn, enables their use in continuously growing cultures. As a proof of concept, we established a sucrose production process that autonomously transitioned from a growth phase with basal production to a production phase with a 5.5-fold higher sucrose titre, triggered by phosphate depletion during cultivation. Overall, this auto-inducible expression system expands the cyanobacterial genetic toolbox and enhances the applicability of cyanobacteria in scalable production processes.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"22 ","pages":"Article e00278"},"PeriodicalIF":4.1,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148157310","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mechanistic and applied study of phosphofructokinases, the “gatekeeper” of the glycolytic pathway on the central carbon metabolism","authors":"Lingyun Li ,&nbsp;Xin Chen ,&nbsp;Yijie Zhang ,&nbsp;Ning Qin ,&nbsp;Yu Chen ,&nbsp;Xu Ji ,&nbsp;Jens Nielsen ,&nbsp;Zihe Liu","doi":"10.1016/j.mec.2025.e00268","DOIUrl":"10.1016/j.mec.2025.e00268","url":null,"abstract":"<div><div>Phosphofructokinase (Pfk), a key regulatory enzyme in glycolysis, is composed of Pfk1 and Pfk2 subunits in <em>Saccharomyces cerevisiae</em>. However, the distinct roles of these subunits in central carbon metabolism remain unclear. Here, we examined the metabolic consequences of deleting <em>PFK1</em> or <em>PFK2</em>. The <em>pfk2Δ</em> strain exhibited more severe defects than <em>pfk1Δ</em>. Its maximum specific growth rate was reduced by approximately 54 % in <em>pfk2Δ</em> and by about 15 % in <em>pfk1Δ</em>, both relative to the reference strain. Ethanol production decreased by 36 % and 82 % in <em>pfk1Δ</em> strain and <em>pfk2Δ</em> strain, respectively, relative to the reference strain. Both deletion strains accumulated higher acetate levels compared to the reference strain, increasing by 25.4 % in the <em>pfk1Δ</em> strain and 82 % in the <em>pfk2Δ</em> strain. Flux balance analysis (FBA) revealed a markedly increased carbon flux to the tricarboxylic acid cycle (TCA) in the <em>pfk2Δ</em> strain, with respiration-associated carbon flux elevated 1.5-fold compared to the <em>pfk1Δ</em> strain. Consistently, transcriptomic profiling showed significant upregulation of respiration-related genes in the <em>pfk2Δ</em> strain compared to the reference strain. Notably, deletion of <em>PFK2</em> enhanced acetyl-CoA-derived product formation, with free fatty acid (FFA) titers increasing from 412 mg L⁻¹ to 517 mg L⁻¹ (a 33.3 % increase). These findings establish <em>PFK2</em> as a key regulatory node redirecting carbon flux from fermentation toward respiration and biosynthesis, offering new opportunities for metabolic engineering of acetyl-CoA-derived products.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"22 ","pages":"Article e00268"},"PeriodicalIF":4.1,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145897977","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Engineering Clostridium thermocellum for production of 2,3-butanediol from cellulose","authors":"S. Bilal Jilani ,&nbsp;Nandhini Ashok ,&nbsp;Yannick J. Bomble ,&nbsp;Adam M. Guss ,&nbsp;Daniel G. Olson","doi":"10.1016/j.mec.2025.e00269","DOIUrl":"10.1016/j.mec.2025.e00269","url":null,"abstract":"<div><div><em>Clostridium thermocellum</em> is a promising host for consolidated bioprocessing due to its ability to directly ferment cellulose into fuels and chemicals. However, natural product formation in this organism is limited. Here, we report engineering <em>C. thermocellum</em> for the production of 2,3-butanediol (23BD), a valuable industrial chemical. We functionally expressed a thermophilic 23BD pathway in this organism resulting in a 23BD titer of 19.7 mM from cellulose, representing a metabolic yield of 24%. We used a cell-free systems biology approach to identify limiting steps in the 23BD pathway, revealing that exogenous 23BD dehydrogenase (BDH) activity was essential for production, while native acetolactate synthase (ALS) and acetolactate decarboxylase (ALDC) activities were present but limiting in the parent strain. This approach also revealed redox balance limitations. We demonstrated that this improved understanding of redox balance limitations could be used to increase 23BD titer in vivo, showing that adding acetate could be used to increase 23BD yield. This work establishes a foundation for developing <em>C. thermocellum</em> into a robust platform for 23BD production directly from cellulose and highlights the utility of cell-free systems for guiding metabolic engineering in non-model organisms.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"22 ","pages":"Article e00269"},"PeriodicalIF":4.1,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145979336","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Elucidating biodegradation of dimethyl terephthalate by two Rhodococcus strains for its valorization applications","authors":"Yifeng Hu ,&nbsp;Tae Seok Moon","doi":"10.1016/j.mec.2026.e00271","DOIUrl":"10.1016/j.mec.2026.e00271","url":null,"abstract":"<div><div>Dimethyl terephthalate (DMT) serves as the precursor in the production of polyethylene terephthalate, polytrimethylene terephthalate, and polybutylene terephthalate. The widespread use of DMT in the polymer industry and its ubiquitous existence in end products raise alarms about its potential harm to humans and animals. DMT can enter the environment through the degradation of polymers and their end products, and cause endocrine disruption, oxidative stress, and an elevated risk of cancer. In recent years, DMT has also gained renewed interest in its potential for plastic recycling and upcycling. In this study, we identified two strains of <em>Rhodococcus</em> that possess DMT-degrading capabilities and utilized transcriptomic analysis and gene knockout to elucidate the mechanisms of DMT degradation. <em>R. opacus</em> PD630 and <em>R. jostii</em> RPET were found to convert up to 1 g/L DMT into mono-methyl terephthalate (MMT). A putative DMTase (RS34275) was identified for this conversion. <em>R. jostii</em> RPET also demonstrates the ability to convert DMT into MMT and to utilize MMT for its cellular growth via the terephthalate pathway. A putative MMTase (RS21885) as the sole enzyme was identified for the conversion of MMT into terephthalate in the RPET strain. In addition, we successfully produced lycopene and lipids from an engineered RPET strain using DMT as a substrate. Our findings will facilitate future DMT bioremediation and bio-upcycling of DMT-associated plastics, enabling the production of value-added products.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"22 ","pages":"Article e00271"},"PeriodicalIF":4.1,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146034885","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A yeast-based platform for etoposide production via yatein bioconversion","authors":"Nicolas Gautron ,&nbsp;Jennifer Perrin ,&nbsp;Ana Luisa Lopez-Vazquez ,&nbsp;Céline Melin ,&nbsp;Marianne Unlubayir ,&nbsp;Killian Tiger ,&nbsp;Cyril Nicolas ,&nbsp;Clément Cuello ,&nbsp;Marc Clastre ,&nbsp;Nicolas Papon ,&nbsp;Nathalie Giglioli-Guivarc’h ,&nbsp;Loïc Guillonneau ,&nbsp;Isabelle Gillaizeau ,&nbsp;Christophe Hano ,&nbsp;Marc Jillian ,&nbsp;Sébastien Besseau ,&nbsp;Vincent Courdavault","doi":"10.1016/j.mec.2026.e00280","DOIUrl":"10.1016/j.mec.2026.e00280","url":null,"abstract":"<div><div>Etoposide, a semisynthetic derivative of podophyllotoxin originally isolated from the rhizomes of mayapple (<em>Podophyllum peltatum</em>) and indian Podophyllum (<em>P. hexandrum</em>) plants, is one of the most powerful chemotherapeutic agents used to treat various types of solid tumors and blood malignancies. Despite its clinical importance, its supply is recurrently constrained due to a heavy reliance on plant extraction, where low natural precursor abundance and increasing climate-related pressures limit production scalability. Developing alternative manufacturing routes has therefore become a major objective, though reconstruction of this complex biosynthetic pathway has long posed significant challenges, even with recent advances in synthetic biology and metabolic engineering. Yeast has emerged as a robust cellular chassis for reconstituting, either partially or entirely, plant secondary metabolite pathways, and enabling cost-effective bioproduction. Here, we established an integrated biotechnological strategy for the sustainable production of advanced etoposide intermediates using engineered yeast cell factories. By combining pathway refactoring, gene copy number optimization, and tailored co-enzyme compatibility, we established an efficient heterologous pathway converting yatein into (−)-4′-desmethyl-epipodophyllotoxin (4′dEPT) in yeast. Iterative strain engineering improved metabolic flux distribution, leading to enhanced titers and accelerated production kinetics, while process engineering proved essential to maximizing overall system performance. Finally, we also demonstrated the viability of coupling bioproduction in cell factories with downstream, semisynthetic conversion by successfully isolating bioreactor-derived 4′dEPT and converting it into etoposide. In parallel, identifying resilient plant resources that can accumulate high levels of YAT provides a complementary strategy for securing the precursor supply at scale. Overall, this report validates the concept of a hybrid etoposide production platform integrating controlled plant biomass sourcing, engineered yeast cell factories, and chemical transformation steps.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"22 ","pages":"Article e00280"},"PeriodicalIF":4.1,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148178464","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metabolic engineering for tobacco quality improvement: Advances and perspectives","authors":"Jie Liu ,&nbsp;Qi Sheng ,&nbsp;Kejuan Zhang ,&nbsp;Jun Zhang ,&nbsp;Xinsheng Li","doi":"10.1016/j.mec.2026.e00281","DOIUrl":"10.1016/j.mec.2026.e00281","url":null,"abstract":"<div><div>As a major cash crop, tobacco quality improvement is constrained by the limited effectiveness of conventional approaches, including traditional breeding and agronomic regulation. This review focuses on the application of metabolic engineering to enable precise and targeted modulation of tobacco quality traits. We first analyze the key factors and underlying mechanisms governing aroma, health-related components, functional metabolites, and combustion characteristics. We then summarize recent advances in metabolic engineering strategies, including pathway rewiring, metabolic flux redistribution, enzyme engineering, and regulatory-network modulation, for optimizing the biosynthesis and accumulation of target compounds such as terpenoids, polyphenols, and alkaloids. In addition, selected integrative approaches that support metabolic regulation and quality formation are briefly discussed. Finally, we highlight current challenges and outline future research directions to facilitate more precise and efficient improvement of tobacco quality.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"22 ","pages":"Article e00281"},"PeriodicalIF":4.1,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148178463","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Substrate stiffness-dependent metabolic reprogramming of iPSC-derived cardiomyocytes on physiological PDMS polymers","authors":"Leena Patel ,&nbsp;Bryan P. Marzullo ,&nbsp;Jonathan Barlow ,&nbsp;Himani Rana ,&nbsp;Amar J. Azad ,&nbsp;Patricia Thomas ,&nbsp;Daniel A. Tennant ,&nbsp;Katja Gehmlich","doi":"10.1016/j.mec.2025.e00266","DOIUrl":"10.1016/j.mec.2025.e00266","url":null,"abstract":"<div><div>Many cardiac pathologies are characterised by increased stiffness of the myocardium, due to excess deposition of extracellular matrix (ECM) proteins and structural remodelling, impacting the behaviour of cardiomyocytes (CMs). Metabolism of CMs shifts in cardiac pathologies, with the healthy heart primarily utilising fatty acids as its source of energy production, whilst the diseased heart switches to utilise glucose. The shift in metabolic source with stiffness of the myocardium has not been investigated.</div><div>To investigate the effect of ECM stiffnesses on iPSC-CM metabolism, iPSC-CMs were cultured on polydimethylsiloxane (PDMS) substrates of healthy and fibrotic stiffness (20 kPa and 130 kPa respectively) and plastic. Cellular metabolism of iPSC-CMs was assessed through isotope-labelled mass spectrometry with central carbon tracing as well as real-time cellular bioenergetics using extracellular flux analysis. Key metabolic genes were investigated at transcript and protein level, with proteomics analysis conducted to identify protein profiles on substrate stiffnesses.</div><div>Mass spectrometry data revealed greater utilisation of glucose in iPSC-CMs cultured on plastic compared to softer PDMS substrates, indicating greater glycolytic activity. Extracellular flux analysis demonstrated greater lactic acid efflux from iPSC-CMs cultured on plastic substrates, reflective of increased glycolytic flux and a shift towards aerobic glycolysis as the primary source of ATP synthesis. This study revealed culture of iPSC-CMs on traditional cell culture plastics or glass coverslips displaying pathological metabolism, highlighting the use of physiological substrates for metabolic investigation.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"21 ","pages":"Article e00266"},"PeriodicalIF":3.7,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144631369","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Model-guided chemical environment and metabolic network design to couple pathways with cell fitness","authors":"Natalia Kakko von Koch ,&nbsp;Tuula Tenkanen ,&nbsp;Sandra Castillo ,&nbsp;Virve Vidgren ,&nbsp;Tino Koponen ,&nbsp;Kristoffer Krogerus ,&nbsp;Merja Penttilä ,&nbsp;Paula Jouhten","doi":"10.1016/j.mec.2025.e00267","DOIUrl":"10.1016/j.mec.2025.e00267","url":null,"abstract":"<div><div>Heterologous compound production is a complex trait since the native metabolic fluxes supplying the precursors, redox power, and energy are under multilevel cellular regulation. Improving complex traits using targeted engineering needs combinatorially charting the complex genetic underpinnings. While this is laborious, adaptive laboratory evolution (ALE) has been used to improve many traits of microbial strains that are of application relevance such as tolerance of harsh conditions and nutrient utilization. However, in contrast to such traits, heterologous production can seldom be intuitively coupled with cellular fitness.</div><div>Here, a novel method EvolveXGA was developed for genome-scale metabolic model guided design of strategies combining chemical environments and genetic engineering of the metabolic network to allow ALE of desired traits. Adaptive evolution of traits occurs when the co-variance between the traits and fitness involves a genetic dependency like a flux coupling would indicate. Thus, combinations of chemical environments and metabolic network structures were searched using a genetic algorithm to identify those that render desired traits (i.e., sets of metabolic fluxes) flux-coupled with fitness. The search was performed for the production of 29 heterologous compounds in yeast <em>Saccharomyces cerevisiae</em>. Strategies for coupling the production routes of 13 compounds with fitness were found with four metabolic reaction knock outs and three components in the chemical environment. In addition, strategies for fitness-coupling native fluxes involved in the production was found for the remaining compounds. In addition, a model-guided strategy was implemented for fitness-coupling of heterologous glycolic acid (GA) synthesis in <em>S</em>. <em>cerevisiae</em> via oxaloacetase, oxalyl-CoA synthetase, and oxalyl-CoA reductase (i.e., oxalate pathway). ALE was performed and evolved populations and isolated clones were characterized using whole-genome sequencing and quantitative metabolite analysis. Three out of six isolates had better GA yield from glucose than a non-optimized control strain expressing the oxalate pathway and glyoxylate reductase.</div><div>EvolveXGA generalizes metabolic model-guided design of strategies to couple production routes with cell fitness. The strategies bring optimizing heterologous production in engineered microbial cells in the realm of ALE. Slow and expensive strain optimization is a major hinder of novel processes using engineered microbial cells reaching industrial realization. Thus, EvolveXGA contributes to biotechnological solutions for the brighter future.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"21 ","pages":"Article e00267"},"PeriodicalIF":4.1,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145620495","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Improvement of D-lactic acid production from methanol by metabolically engineered Komagataella phaffii via ultra-violet mutagenesis","authors":"Yoshifumi Inoue,&nbsp;Kaito Nakamura,&nbsp;Ryosuke Yamada,&nbsp;Takuya Matsumoto,&nbsp;Hiroyasu Ogino","doi":"10.1016/j.mec.2025.e00262","DOIUrl":"10.1016/j.mec.2025.e00262","url":null,"abstract":"<div><div>Methanol has attracted attention as an alternative carbon source to petroleum. <em>Komagataella phaffii</em>, a methanol-assimilating yeast, is a useful host for the chemical production from methanol. A previous study successfully constructed a metabolically engineered <em>K. phaffii</em> GS115/S8/Z3 strain capable of producing D-lactic acid from methanol. In this study, we aimed to develop a strain with improved D-lactic acid production by applying ultra-violet mutagenesis to the D-lactic acid-producing strain, GS115/S8/Z3. The resulting mutant strain DLac_Mut2_221 produced 5.38 g/L of D-lactic acid from methanol, a 1.52-fold increase compared to the parent strain GS115/S8/Z3. The transcriptome analysis of the mutant DLac_Mut2_221 identified 158 differentially expressed genes, providing insights into key mechanisms contributing to enhanced D-lactic acid production. Metabolic engineering strategies for <em>K. phaffii</em> based on the knowledge gained from this study will contribute to improving the productivity of various useful chemicals from methanol.</div></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":"20 ","pages":"Article e00262"},"PeriodicalIF":3.7,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144089620","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}