Microbial Cell Factories最新文献

{"title":"Genomically integrated orthogonal translation system in Escherichia coli enables production of functional modified [NiFe]-hydrogenases.","authors":"Qin Fan, Stevanie Stevanie, Stefan Frielingsdorf, Peter Neubauer, Oliver Lenz, Matthias Gimpel","doi":"10.1186/s12934-026-03057-z","DOIUrl":"10.1186/s12934-026-03057-z","url":null,"abstract":"<p><p>The functional diversification of O₂-tolerant [NiFe]-hydrogenases using orthogonal translation systems (OTSs) offers a promising strategy for developing advanced biocatalysts and biohybrid energy platforms. However, plasmid-based OTSs frequently impose metabolic burdens and suffer from plasmid instability during fermentation, particularly when co-produced with complex metalloenzymes. To overcome these bioprocess limitations, we employed CRISPR/Cas9-mediated genome editing to integrate a psychrophilic pyrrolysyl-tRNA synthetase/tRNA pair into the Escherichia coli BL21 genome. The resulting strain provided a plasmid-free orthogonal translation background that supported amber suppression-mediated expression of the regulatory [NiFe]-hydrogenase (RH) of Cupriavidus necator. Using this genomically integrated OTS, we achieved the production of a full-length, catalytically active RH variant. Our results demonstrate that chromosomal OTS is compatible with the efficient production and maturation of complex metalloenzymes. The present work lays the groundwork for the bio-orthogonal engineering of hydrogenases and related hybrid biocatalysts.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-07-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13340272/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148397181","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The transcriptional response to environmental alkalization involves both Komagataella phaffii Pho4 transcription factors.","authors":"Marcel Albacar, Antonio Casamayor, Rui Wang, Joaquín Ariño, Asier González","doi":"10.1186/s12934-026-03054-2","DOIUrl":"10.1186/s12934-026-03054-2","url":null,"abstract":"<p><strong>Background: </strong>Pho4 is a fungal transcription factor essential for the response to phosphate (Pi) starvation in many fungi and is usually encoded by a single gene. However, we recently reported that the methylotrophic yeast Komagataella phaffii encodes two functional Pho4 proteins, named Pho4(A) and Pho4(B), and that only Pho4(B) was required for the transcriptional response to Pi limitation. Pho4 is also important in many yeasts for adaptation to an alkaline environment. We evaluate here the participation of Pho4(A) and Pho4(B) in the transcriptional response to alkaline pH.</p><p><strong>Results: </strong>We show that the expression of nearly one-hundred genes is altered in a pho4(A) pho(B) mutant. In contrast to Pi starvation, Pho4(A) becomes important under high pH stress for the short-time expression of genes required for Pi homeostasis, such as PHO89, VTC1, VTC2, VTC4 and PHO4(B), suggesting that both transcription factors respond to different signals. Integration of the transcriptional data with that obtained from crz1 or rim101 mutants shows that nearly 80 genes are regulated by at least two of these transcription factors. Thus, we show that induction of PHO89 by Pi starvation or high pH is fully abolished in both double pho4(A) pho4(B) and single rim101 mutants, and that mutation of CRZ1 and RIM101 partially eliminates the sensitivity to high pH of the pho4(A) pho4(B) strain, revealing unexpected genetic interactions between these mutations. Finally, we identify an alkali-sensitive region in the PHO89 promoter and demonstrate that its duplication further increases the promoter's response to alkalinization.</p><p><strong>Conclusions: </strong>The transcription factor Pho4(A) is not required for the transcriptional response to Pi starvation, but it contributes to the response to alkalinization. Gene expression remodeling upon alkalinization in K. phaffii is controlled by diverse signaling pathways, including Pho4, Crz1 and Rim101 as well as other components still to uncover. Detailed knowledge of the signaling mechanisms triggered upon alkalinization and promoter mapping of the genes responsive to high pH will be useful tools for the generation of alkaline pH-regulated hybrid synthetic promoters in K. phaffii with improved features.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13613856/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148388098","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Acetate as alternative carbon source for production of mono- and di-rhamnolipids in Pseudomonas putida KT2440.","authors":"Jakob Grether, Sarah Leibinger, Christina Ramke, Philipp Hubel, Lisa Weber, Jens Pfannstiel, Elvio Henrique Benatto Perino, Rudolf Hausmann","doi":"10.1186/s12934-026-03050-6","DOIUrl":"10.1186/s12934-026-03050-6","url":null,"abstract":"<p><strong>Background: </strong>The bacterium Pseudomonas putida KT2440 is used as a safe platform for rhamnolipid production via expression of the rhlABC genes, yielding mono- and di-rhamnolipid congeners. While glucose is commonly used as a carbon source, more sustainable alternatives such as acetate from waste streams or electrolysis are gaining interest. These two carbon sources are assimilated into the core carbon metabolism through distinct entry points. The impact of this on the proteome and rhamnolipid congener composition remain unclear. In this study, P. putida JAG1 was used to compare rhamnolipid production and composition during growth on glucose and acetate. Additionally, the impact of formate as a non-assimilable electron donor and the potential of modeled electrochemically derived CO₂-based carbon mixtures were evaluated.</p><p><strong>Results: </strong>Rhamnolipid precursor synthesis involves the rhamnose pathway and de-novo fatty acid synthesis. While glucose feeds directly into the rhamnose pathway and acetate into fatty acid synthesis, both carbon sources resulted in a similar di-rhamnolipid fraction of approximately 75 mol%. Although the carbon source significantly influenced the proteome profile, the carbon-to-nitrogen consumption ratio remained constant (C/N ≈ 15). Proteomic differences in P. putida JAG1 comparing the two carbon sources suggest ATP independent acetyl-CoA recycling as energy saving strategy. Enzyme abundances moreover suggest a coupling of the glyoxylate shunt and glycerol metabolism as strategy for gluconeogenesis. Growth rates, as well as biomass and product yields, were slightly lower on acetate compared to glucose. Co-feeding formate with acetate did not cause significant changes relative to acetate alone. Simultaneous consumption of acetate, formate, and ethanol only slightly affect product-per-biomass yields but strongly shifted the composition toward mono-rhamnolipids. Furthermore, comparison of the di-rhamnolipid producer with a mono-rhamnolipid producer indicates that di-rhamnolipid formation is associated with a higher molar product yield.</p><p><strong>Conclusions: </strong>Acetate resembles a viable alternative carbon source for rhamnolipid production with only slight drawbacks to glucose. The metabolism of the microbial cell factory P. putida JAG1 appears homeostatic in presence of a single assimilable carbon source without affecting the rhamnolipid congener composition. The availability of a second assimilable carbon source, however, shifted the composition in favor of the mono-rhamnolipid. The molar product yield per carbon atom was higher during production of di-rhamnolipids compared to sole production of mono-rhamnolipids, highlighting it as a promising target product for future studies.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":"25 1","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-07-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13330470/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148376194","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cell-free synthesis and characterization of Salmonella, Escherichia coli, and Shigella-specific bacteriophages.","authors":"Irem Iskender, Jaap Bosma, Paul Soudier, Yating Guo, Jinquan Li, Vincent Noireaux, Steven Bowden","doi":"10.1186/s12934-026-03049-z","DOIUrl":"10.1186/s12934-026-03049-z","url":null,"abstract":"<p><strong>Background: </strong>Cell-free gene expression (CFE) systems provide a rapid and modular platform for synthesizing bacteriophages without the need for living host cells. However, the generalizability of CFE-based phage synthesis across diverse phages, as well as the functional comparability of in vitro-synthesized phages to host-derived counterparts, has not yet been fully explored. In this study, we evaluated the ability of an Escherichia coli-based CFE platform to synthesize diverse bacteriophages relevant to food safety and synthetic biology applications.</p><p><strong>Results: </strong>Using an E. coli-based CFE system, we synthesized seven phages from four different families, achieving infectious titers ranging from 10⁶ to 10¹¹ plaque-forming units per milliliter (PFU/mL). Of these seven phages, five phages, vB_SalM-LPST153 (LPST153), vB_SenM-S16 (S16), SP6, vB_Sens_Jbel (Jbel), and vB_EcoM_Alf5 (Alf5), are reported here for the first time as successfully synthesized and characterized in a CFE system. CFE-synthesized phages remained capable of infecting host strains; however, significant differences in efficiency of plating (EOP) were observed across multiple Salmonella, E. coli, and Shigella hosts. Long-term storage tests for LPST153 showed that phage buffer (PB) stably maintained CFE titers above 10⁹ PFU/mL for 12 weeks, while purified CFE-synthesized and host-derived LPST153 lysates exhibited broadly comparable thermal stability profiles across the tested temperatures. Using the PHEIGES workflow, we also demonstrated complete in vitro assembly and expression of LPST153 from PCR fragments. In addition, phages were successfully synthesized using CFE lysates derived from the endotoxin-free E. coli strain ClearColi™, demonstrating the synthesis of infectious phages in an endotoxin-free system.</p><p><strong>Conclusions: </strong>These findings expand the diversity of bacteriophages compatible with cell-free synthesis (CFS) and demonstrate that CFE-synthesized phages remain infectious and retain several functional properties, although some differences in host-dependent infectivity were observed. The ability to synthesize infectious phages in endotoxin-free systems and assemble phage genomes entirely in vitro highlights the potential of CFE platforms for scalable, decentralized, and cold-chain-independent manufacturing of phage-based therapeutics and food safety interventions.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-06-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13591810/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148361595","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Correction: Bacillus thuringiensis as a factory producing parasporin toxins with promising anticancer properties: a review.","authors":"Natalia Musiuk, Wioleta Lewandowska, Aleksandra Rembiszewska, Izabela Święcicka","doi":"10.1186/s12934-026-03046-2","DOIUrl":"10.1186/s12934-026-03046-2","url":null,"abstract":"","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":"25 1","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13292566/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148318203","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multi-omics-guided metabolic engineering of Limosilactobacillus reuteri for high-level 3-hydroxypropionaldehyde production from glucose.","authors":"Zuojun Liu, Qiang Yin, Liang Li, Hang Wu, Yong Fang, Fuqiang Cheng, Ting Guo","doi":"10.1186/s12934-026-03048-0","DOIUrl":"10.1186/s12934-026-03048-0","url":null,"abstract":"<p><strong>Background: </strong>Limosilactobacillus reuteri is an important probiotic chassis for producing 3-hydroxypropionaldehyde (3-HPA), a broad-spectrum antimicrobial and biochemical. However, its production is constrained by a metabolic trilemma comprising coenzyme B₁₂ auxotrophy, redox imbalance, and carbon catabolite repression (CCR). The main bottleneck currently limiting cost-effective glucose-based production is the inability to simultaneously resolve these interconnected constraints.</p><p><strong>Results: </strong>Through integrated multi-omics analyses of differential carbon source performance, an unexpected regulatory mechanism was obtained. Galactose orchestrates a tripartite metabolic program in which UDP-galactose may act as a signaling molecule potentially involved in alleviating CCR; redirects carbon flux through the pentose phosphate pathway to balance ATP/NADH, and transcriptionally co-activates dhaB (glycerol dehydratase) and cobA/Q (B₁₂ biosynthesis) for coordinated holoenzyme assembly. Strikingly, this native regulatory network enables 8.51 g/L 3-HPA from galactose despite its poor support for cell growth, versus 1.67 g/L from glucose which readily supports robust growth-a 5.1-fold advantage that highlights galactose's metabolic priming efficacy. Guided by these insights, an engineered strain overexpressing glycerol dehydratase was combined with process optimization and a rationally designed four-factor targeted supplementation. Synergistic supplementation with coenzyme B₁₂, ATP, KCl, and MgCl₂ achieved 28.96 g/L 3-HPA-a 17.34-fold improvement over native glucose-based performance.</p><p><strong>Conclusions: </strong>To our knowledge, this is the first systems-level strategy for compensating the metabolic trilemma in L. reuteri by engineering carbon source-dependent regulatory networks and cofactor dynamics in a cost-effective glucose-based system, achieving the highest 3-HPA titer reported in lactobacilli in shake-flask fermentation to date. This work establishes a scalable platform for industrial 3-HPA production and provides a systems-level framework for reprogramming central carbon metabolism in probiotic chassis to address persistent bioproduction bottlenecks.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-06-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13539820/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148302296","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Exploring the metabolic burden of surfactin biosynthesis and the metabolic costs of srfA operon expression in Bacillus subtilis.","authors":"Elvio Henrique Benatto Perino, Eric Hiller, Maliheh Vahidinasab, Sandra Moneta, Bahar Abrishamchi, Kowsala Nagendran, Philipp Hubel, Jens Pfannstiel, Rudolf Hausmann","doi":"10.1186/s12934-026-03047-1","DOIUrl":"10.1186/s12934-026-03047-1","url":null,"abstract":"<p><p>The biosynthesis of surfactin, a potent lipopeptide biosurfactant produced by Bacillus subtilis, imposes a significant metabolic burden on the host organism. Understanding the metabolic costs associated with surfactin production, specifically the ATP demand and precursor diversion resulting from the expression of the large srfAA-AD operon (srfA operon), is crucial for optimizing production strains. In this study, the metabolic burden and growth impacts associated with surfactin biosynthesis in B. subtilis were quantitatively evaluated by comparing a reference surfactin-producing strain (BMV9) with two mutant strains: BMV12, which lacks the srfA operon, and BMV33, which retains the srfA operon but lacks the sfp gene. Our analysis included theoretical calculations of ATP and NADPH + H⁺ requirements for de novo surfactin synthesis, and we measured growth behavior and carbon and nitrogen source consumption. Results indicated that surfactin production significantly reduces biomass yield (Y_X/S) and specific growth rates due to the metabolic costs of expressing the non-ribosomal peptide synthetase (NRPS) and the diversion of key precursors. Notably, BMV12 exhibited higher growth rates compared to the surfactin-producing strain. Proteome analyses further revealed differential protein abundance in non-surfactin-producing strains, indicating altered metabolic pathways that may relieve the metabolic burden associated with surfactin synthesis. These findings highlight the complex trade-offs between secondary metabolite production and cellular growth, providing a foundation for metabolic engineering strategies aimed at optimizing surfactin yields while minimizing metabolic costs.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-06-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13289377/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148283973","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Biological production of muconic acid from renewable biomass: advances in metabolic engineering and lignin valorisation.","authors":"Vinod Kumar, Satish Kumar Ainala, Vivek Kumar Gaur, Samuel Jacob, Yuheng Lin, Ponnusami Venkatachalam, Sunil K Maity, Naseem A Gaur, Gopalakrishnan Kumar, Vijai Kumar Gupta","doi":"10.1186/s12934-026-03037-3","DOIUrl":"https://doi.org/10.1186/s12934-026-03037-3","url":null,"abstract":"<p><p>Muconic acid (MA) is characterized by two reactive carboxylic acid groups and two conjugated double bonds, making it a highly valuable industrial platform chemical with significant market potential. It serves as a key intermediate in the manufacturing of important commercial chemical products such as adipic acid and terephthalic acid. The finite fossil-based resources and climate issues due to CO₂ emission have necessitated the microbial routes for the production of MA, a potential alternative to fossil-based synthesis. This review provides a comprehensive overview of recent progress in the biological production of MA. The article begins with an outline of the present catalytic routes and known biochemical pathways for MA biosynthesis. The review then focuses on metabolic engineering strategies employed in various microbial hosts including Escherichia coli, Corynebacterium glutamicum, and Pseudomonas putida to enhance MA production from diverse feedstocks such as sugars, aromatic compounds, and lignin-derived substrates. Special attention is given to pathway optimization, host tolerance, and strategies enabling efficient conversion of lignin-derived intermediates within integrated biorefinery frameworks. Key challenges associated with scaling up bio-based MA production to industrial levels are discussed, along with potential strategies for developing robust and efficient microbial cell factories. The review concludes with future perspectives and recommendations to accelerate research progress and development in this field.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":4.9,"publicationDate":"2026-06-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148258817","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification and characterization of a novel leaderless bacteriocin toyoncin T35 from Bacillus toyonensis.","authors":"Xiaofeng Zhang, Haihua Wang, Chenxu He, Xiaojing Zhang, Xuesi Li, Chunmei Pan","doi":"10.1186/s12934-026-03044-4","DOIUrl":"10.1186/s12934-026-03044-4","url":null,"abstract":"<p><strong>Background: </strong>Antibiotics residues and antibiotics resistance have become a serious threat to human health. There is an urgent need for the development of antibiotic alternatives. Bacteriocins are considered one of the alternative to antibiotics because of their different antimicrobial mechanisms from antibiotics. Therefore, mining novel bacteriocins is of great significance for the development of antibiotic alternatives.</p><p><strong>Results: </strong>A novel aureocin A53-like leaderless bacteriocin, toyoncin T35, was identified in B. toyonensis T35. Toyoncin T35 was composed of 48 amino acid residues and its molecular weight was 5491.9950 Da. Toyoncin T35 exhibited strong antimicrobial activity against foodborne pathogens B. cereus (MIC, 0.71 µM), L. monocytogenes (MIC, 0.35 µM) as well as pathogenic bacteria S. suis (MIC, 2.84 µM). In addition, toyoncin T35 could inhibit B. cereus spore outgrowth, and possessed broad acid-base adaptability (pH 2-10) along with high heat tolerance (below 100 °C). Toyoncin T35 has no hemolysis activity and cytotoxicity even at a high concentration of 91.00 µM.</p><p><strong>Conclusions: </strong>The novel leaderless bacteriocin toyoncin T35 has the potential to become an antibiotic alternative for treating diseases caused by foodborne pathogens B. cereus, L. monocytogenes, and pathogenic bacteria S. suis.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-06-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13491596/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148252290","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metabolomic insights into functional protein mediated salt stress adaptation in the osmophilic yeast Zygosaccharomyces rouxii.","authors":"Dingkang Wang, Lerong Liu, Sha Liu, Ting Shi, Yihe Bu, Yanqin Rong, Shiyu Liu, Longzhan Gan, Yiting Kang, Yongjie Zhang","doi":"10.1186/s12934-026-03018-6","DOIUrl":"10.1186/s12934-026-03018-6","url":null,"abstract":"<p><strong>Background: </strong>High salinity inhibits microbial growth by disturbing osmotic balance, redox equilibrium, and membrane integrity. Zygosaccharomyces rouxii, an osmophilic yeast widely used in high-salt fermented foods, has evolved adaptive responses to such stress, but the global metabolic mechanisms underlying these responses, especially under functional protein supplementation, remain poorly understood.</p><p><strong>Methods and results: </strong>In this study, an untargeted UPLC-HRMS-based metabolomics approach was used to investigate the metabolic responses of Z. rouxii to high-salt stress and functional protein supplementation. Multivariate analysis revealed significant metabolic reprogramming at 120 g/L NaCl, while functional protein supplementation induced a distinct metabolic state beyond that caused by salt stress alone. Pathway enrichment analysis showed that protein supplementation markedly enhanced amino acid metabolism, antioxidant-related pathways, and nutrient transport, while preferentially modulating lipid metabolism associated with membrane remodeling. In particular, pathways related to unsaturated fatty acids and glycerolipids were finely regulated, suggesting improved membrane flexibility under hyperosmotic conditions.</p><p><strong>Conclusions: </strong>Collectively, these findings indicate that functional protein supplementation enhances coordinated metabolic regulation in Z. rouxii, thereby improving metabolic flexibility and salt stress tolerance. This study provides mechanistic insights into osmotic stress adaptation and proposes a feasible strategy to enhance yeast robustness under high-salt conditions.</p>","PeriodicalId":18582,"journal":{"name":"Microbial Cell Factories","volume":" ","pages":""},"PeriodicalIF":5.8,"publicationDate":"2026-06-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13491812/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148252287","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}