Synthetic and Systems Biotechnology最新文献

{"title":"Promoter engineering for enhanced 3-hydroxypropionic acid production in Pichia pastoris","authors":"Shushu Chen ,&nbsp;Mengyao Zhang ,&nbsp;Xiaoyan Wu ,&nbsp;Fan Bai ,&nbsp;Linhui Gao ,&nbsp;Yiwei Shen ,&nbsp;Shaohua Dou ,&nbsp;Peng Cai ,&nbsp;Yongjin J. Zhou","doi":"10.1016/j.synbio.2025.04.013","DOIUrl":"10.1016/j.synbio.2025.04.013","url":null,"abstract":"<div><div>Enabling tools are essential for facilitating the methanol bioconversion in <em>Pichia pastoris</em>. However, there is still a relative lack of promoters that can stably express high levels without being affected by the carbon source, which hinders the construction and modification of cell factories containing long metabolic pathways. This study mapped a gene expression intensity library of central metabolic pathways in <em>P. pastoris</em> under methanol and glucose conditions. Through modification of the upstream sequences of promoters, an artificial promoter P_<em>S2</em> was developed with a strong intensity up to 90 % of P_<em>GAP</em>. By using this promoter, we successfully constructed a hybrid pathway that integrates the β-alanine and malonyl-CoA pathways for the production of 3-hydroxypropionic acid. Further combining rational metabolic engineering strategies, such as optimizing gene copy numbers and blocking byproduct synthesis pathways, the engineered strains CHP9 and CHP20 achieved 3-HP titers of 23 g/L and 22 g/L by using methanol as the sole carbon source. These results indicate that adaptive strength of promoters can facilitate efficient chemical biosynthesis in methanol bioconversion by mitigating glucose repression effects. This work preliminarily explored the expression patterns of genes in the central metabolic pathways of <em>P. pastoris</em>, identified and characterized the intensities of various endogenous promoters, and extended the enabling toolbox for <em>P. pastoris</em>. This result also lays a foundation for the construction of microbial cell factories and the industrial production of 3-HP via methanol bioconversion.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 916-924"},"PeriodicalIF":4.4,"publicationDate":"2025-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143899444","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":"Ty retrotransposon element based multiple integration toolkit for Saccharomyces cerevisiae","authors":"Song Gao ,&nbsp;Weizhu Zeng ,&nbsp;Dong Li ,&nbsp;Sha Xu ,&nbsp;Jingwen Zhou","doi":"10.1016/j.synbio.2025.04.011","DOIUrl":"10.1016/j.synbio.2025.04.011","url":null,"abstract":"<div><div>Extra-high-level overexpression of single or multiple specific proteins by integrating specific genes in the genome is vital to achieve the stable and efficient production of target proteins and metabolites in <em>S. cerevisiae</em>. Five families of Ty elements in the genome of <em>S. cerevisiae</em> CEN.PK2-1D, which could have dozens to hundreds of copies, have been employed to achieve massive gene expression. By engineering nine selective markers, six of them (<em>TRP1</em>, <em>LEU2</em>, <em>URA3</em>, <em>HIS5</em>, <em>natMX</em> and <em>hphMX</em>) achieve stably high copy integration (&gt;15 copies) at Ty sites. Fluorescence proteins and taxifolin biosynthesis pathway genes were overexpressed to verify the toolkit. The titer of protein phiYFP in the multiple integration strain reached 1.6 g/L (268.1 mg/g DCW), and its fluorescence intensity was 3.3 times higher than that in the episomal overexpression strain. For taxifolin biosynthesis, 14 genes were integrated into three different Ty sites using three selective markers from the toolkit, resulting in 277.6 mg/L taxifolin accumulation from glucose.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 887-896"},"PeriodicalIF":4.4,"publicationDate":"2025-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143887471","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":"Extending dynamic and operational range of the biosensor responding to l-carnitine by directed evolution","authors":"Tingting Li ,&nbsp;Huina Dong ,&nbsp;Jinlong Li ,&nbsp;Huiying Wang ,&nbsp;Chunxiang Pu ,&nbsp;Siyu Chen ,&nbsp;Zhiying Yang ,&nbsp;Xinyi Ren ,&nbsp;Xuan Liu ,&nbsp;Zhaoxia Jin ,&nbsp;Dawei Zhang","doi":"10.1016/j.synbio.2025.04.012","DOIUrl":"10.1016/j.synbio.2025.04.012","url":null,"abstract":"<div><div><span>l</span>-carnitine is a quaternary amine compound essential for eukaryotic metabolism. It is mainly involved in the oxidative decomposition of medium-and long-chain fatty acids and provides energy for the body. Therefore, it is widely used in health care and food additives. As a pivotal transcriptional activator of <span>l</span>-carnitine metabolism, CaiF is notably activated by crotonobetainyl-CoA, a key intermediate product in the carnitine metabolic pathway. Capitalizing on this mechanism, a sophisticated biosensor was ingeniously developed. Nevertheless, it is worth mentioning that the biosensor currently exhibits a relatively restricted detection range, which results in some specific limitations in practical application scenarios. In this paper, we constructed a biosensor based on CaiF and developed a strategy for modifying this biosensor. The structural configuration of CaiF was formulated by computer-aided design, and the DNA binding site was simulated, which was verified by alanine scanning. Functional Diversity-Oriented Volume-Conservative Substitution Strategy of the key sites of CaiF was conducted to extend the dynamic range of the biosensor. The biosensor based on CaiF^Y47W/R89A, which exhibited a considerably expanded concentration response range, from 10⁻⁴ mM–10 mM, was obtained. The response range was 1000-fold wider and the output signal intensity was 3.3-fold higher to that of the control biosensor. These variants may have great value in improving the <span>l</span>-carnitine production process.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 897-906"},"PeriodicalIF":4.4,"publicationDate":"2025-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143886845","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":"Sequence and taxonomic feature evaluation facilitated the discovery of alcohol oxidases","authors":"Yilei Han ,&nbsp;Xuwei Ding ,&nbsp;Junjian Tan ,&nbsp;Yajuan Sun ,&nbsp;Yunjiang Duan ,&nbsp;Zheng Liu ,&nbsp;Gaowei Zheng ,&nbsp;Diannan Lu","doi":"10.1016/j.synbio.2025.04.014","DOIUrl":"10.1016/j.synbio.2025.04.014","url":null,"abstract":"<div><div>Recent advancements in data technology offer immense opportunities for the discovery and development of new enzymes for the green synthesis of chemicals. Current protein databases predominantly prioritize overall sequence matches. The multi-scale features underpinning catalytic mechanisms and processes, which are scattered across various data sources, have not been sufficiently integrated to be effectively utilized in enzyme mining. In this study, we developed a sequence- and taxonomic-feature evaluation driven workflow to discover enzymes that can be expressed in <em>E. coli</em> and catalyze chemical reactions <em>in vitro</em>, using alcohol oxidase (AOX) for demonstration, which catalyzes the conversion of methanol to formaldehyde. A dataset of 21 reported AOXs was used to construct sequence scoring rules based on features, including sequence length, structural motifs, catalytic-related residues, binding residues, and overall structure. These scoring rules were applied to filter the results from HMM-based searches, yielding 357 candidate sequences of eukaryotic origin, which were categorized into six classes at 85 % sequence similarity. Experimental validation was conducted in two rounds on 31 selected sequences representing all classes. Among these selected sequences, 19 were expressed as soluble proteins in <em>E. coli</em>, and 18 of these soluble proteins exhibited AOX activity, as predicted. Notably, the most active recombinant AOX exhibited an activity of 8.65 ± 0.29 U/mg, approaching the highest activity of native eukaryotic enzymes. Compared to the established UniProt-annotation-based workflow, this feature-evaluation-based approach yielded a higher probability of highly active recombinant AOX (from 8.3 % to 19.4 %), demonstrating the efficiency and potential of this multi-dimensional feature evaluation method in accelerating the discovery of active enzymes.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 907-915"},"PeriodicalIF":4.4,"publicationDate":"2025-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143886847","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":"T7 RNA polymerase-guided base editor for accelerated continuous evolution in Bacillus subtilis","authors":"Bin Wang ,&nbsp;Yaokang Wu ,&nbsp;Xueqin Lv ,&nbsp;Long Liu ,&nbsp;Jianghua Li ,&nbsp;Guocheng Du ,&nbsp;Jian Chen ,&nbsp;Yanfeng Liu","doi":"10.1016/j.synbio.2025.04.010","DOIUrl":"10.1016/j.synbio.2025.04.010","url":null,"abstract":"<div><div>Targeted <em>in vivo</em> hypermutation mediated by base deaminase-T7 RNA polymerase (T7 RNAP) fusions promotes genetic diversification and accelerates continuous directed evolution. Due to the lack of a T7RNAP expression regulation system and functionally compatible linker for fusion protein expression, T7RNAP-guided continuous evolution has not been established in <em>Bacillus subtilis</em>, which limited long gene fragment continuous evolution targeted on genome. Here, we developed BS-MutaT7 system, which introduced mutations into specific genomic regions by leveraging chimeric fusions of base deaminases with T7RNAP in <em>B. subtilis.</em> We selected seven different sources of adenosine and cytosine deaminases, 14 fusion protein linkers to be fused with T7RNAP, constructing four libraries with the size of 5000, where deaminases were fused at either the N- or C-terminus of T7RNAP. Based on the efficiency of binding to T7 promoter and high mutagenesis activity, two optimal chimeric mutators, BS-MutaT7^A (TadA8e-Linker0-T7RNAP) and BS-MutaT7^C (PmCDA1-(GGGGS)₃-T7RNAP co-expressed with UGI) were identified. The target mutation rates reached 1.2 × 10⁻⁵ per base per generation (s.p.b.) and 5.8 × 10⁻⁵ s.p.b., representing 7000-fold and 37,000-fold increases over the genomic mutation rate, respectively. Both exhibited high processivity, maintaining mutation rates of 5.8 × 10⁻⁶ s.p.b. and 2.9 × 10⁻⁵ s.p.b. within a 5 kb DNA region. Notably, BS-MutaT7^C exhibited superior mutagenic activity, making it well-suited for applications requiring intensive and sustained genomic diversification. Application of BS-MutaT7 enabled a 16-fold increase in tigecycline resistance and enhanced β-lactoglobulin (β-Lg) expression by evolving the global transcriptional regulator <em>codY</em>, achieving a β-Lg titer of 3.92 g/L. These results highlight BS-MutaT7 as a powerful and versatile tool for genome-scale continuous evolution in <em>B. subtilis</em>.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 876-886"},"PeriodicalIF":4.4,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143887470","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":"Integration of metabolic and evolutionary processes to construct efficient xylose-utilizing strain of Zymomonas mobilis for lignocellulosic ethanol production","authors":"Jiyun Lou ,&nbsp;Xia Wang ,&nbsp;Runxia Li ,&nbsp;Jun Yao ,&nbsp;Junpeng Hu ,&nbsp;Xianwu Qin ,&nbsp;Mingjie Jin ,&nbsp;Shihui Yang","doi":"10.1016/j.synbio.2025.04.009","DOIUrl":"10.1016/j.synbio.2025.04.009","url":null,"abstract":"<div><div>Xylose is a major component of lignocellulose, and it is crucial to enable microorganisms the capability of efficient xylose utilization for economical lignocellulosic biochemical production. <em>Zymomonas mobilis</em> is a natural ethanologenic bacterium with xylose-utilizing recombinant strains engineered for commercial lignocellulosic ethanol production. However, low efficiency of xylose utilization is still the major hurdle for lignocellulosic biochemical production, especially in the lignocellulosic hydrolysate containing inhibitory compounds and mixed sugars of glucose and xylose. In this study, xylose-utilization capability of a xylose-utilizing recombinant strain 8b-S38 of <em>Z. mobilis</em> was further improved by evaluating the effects of different xylose metabolic pathways, xylose isomerases, and xylose transporters. The results demonstrated that xylose isomerase pathway is still the most efficient one in <em>Z. mobilis</em> among the xylose metabolic pathways examined, and the introduction of xylose isomerase (XI) such as StrXI and PanXI to increase its copy numbers can improve xylose utilization. In addition, the introduction of effective xylose transporter such as high-affinity xylose transporter XylFGH and the glucose facilitator mutant Glf^{A165M−K458I} promoted xylose transportation and subsequent utilization. A recombinant strain S67SPGlf^m was constructed combining rational engineering of xylose isomerases and transporters and semi-rational adaptation, which exhibited enhanced xylose utilization capability in lignocellulosic hydrolysate. This study illustrated that the construction of robust and efficient industrial strains with complex phenotypes requires a combination of different strategies such as metabolic engineering and adaptive laboratory evolution, which also provided biological parts such as xylose isomerases and xylose transporters to help design and construct microbial cell factories for efficient xylose utilization in the future.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 858-867"},"PeriodicalIF":4.4,"publicationDate":"2025-04-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143869540","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":"Nonglycosidic C–O bond formation catalyzed by a bifunctional pseudoglycosyltransferase ValL","authors":"Ziyue Guo ,&nbsp;Xin Zhang ,&nbsp;Lin Zhou ,&nbsp;Qungang Huang ,&nbsp;Qianjin Kang ,&nbsp;Linquan Bai","doi":"10.1016/j.synbio.2025.04.007","DOIUrl":"10.1016/j.synbio.2025.04.007","url":null,"abstract":"<div><div>The C₇N antibiotic validamycin A is an antifungal agent widely used as a crop protectant. It comprises a validoxylamine A unit linked to a glucose moiety, which is formed through a nonglycosidic <em>C</em><em>–</em><em>N</em> bond connecting a valienol moiety and a validamine moiety, a reaction catalyzed by the pseudoglycosyltransferase ValL. In this study, we analyzed the chemical composition of validamycins in <em>Streptomyces hygroscopicus</em> var. <em>jinggangensis</em> TL01. A series of novel oxygen-bridged analogues, namely, validenomycin, validomycin, and 1,1′-bis-valienol, were identified in the culture supernatants, and their chemical structures were elucidated using a combination of one- and two-dimensional nuclear magnetic resonance and mass spectrometry. Gene disruption and complementation experiments revealed that <em>valL</em> is essential for the biosynthesis of these new oxygen-bridged analogues of validamycins. Biochemical assays further demonstrated that ValL catalyzed the <em>C–O</em> bond formation between GDP-valienol and valienol-7-phosphate, producing 1,1′-bis-valienol-7-phosphate, which was subsequently dephosphorylated by ValO and glycosylated by ValG to yield validenomycin. Collectively, our findings revealed the unique ability of ValL to catalyze nonglycosidic <em>C–O</em> coupling, potentially enabling the generation of various chemical scaffolds for C₇N family antibiotics.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 846-857"},"PeriodicalIF":4.4,"publicationDate":"2025-04-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143864111","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":"Network of nicotinamide mononucleotide biosynthesis: substrate tracking, rate-limiting enzyme, and regulatory strategy","authors":"Puyue Zhang ,&nbsp;Ziru Ye ,&nbsp;Zhong Tian ,&nbsp;Qing Liu ,&nbsp;Yong Huang","doi":"10.1016/j.synbio.2025.04.005","DOIUrl":"10.1016/j.synbio.2025.04.005","url":null,"abstract":"<div><div>Nicotinamide mononucleotide (NMN) is a nucleotide of significant biological importance, found abundantly in various foods such as meat, fruits, and vegetables. Recently, its potential effects in delaying aging have attracted considerable attention. Although chemical synthesis methods are commonly employed, they do not align with green production standards. In contrast, the biosynthesis of NMN is both safer and more environmentally sustainable. In this review, we established a novel “substrate-pathway-enzymology” framework to analyze the research on NMN biosynthesis. First, we systematically trace four substrates (nicotinamide ribose, nicotinamide, niacin, and nicotinamide adenine dinucleotide) and their respective metabolic routes. Then, we thoroughly investigate key enzymes through structural biology and protein engineering approaches, and converge the fragmented research findings across pathways to construct a comprehensive NMN biosynthesis network, revealing intricate metabolic regulations and pathway interactions. Through comparative analysis, the most promising biosynthetic pathway and prospects are discussed. Additionally, this review also provides original perspectives for NMN industrial development.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 959-972"},"PeriodicalIF":4.4,"publicationDate":"2025-04-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144068098","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":"A systematic study of regulating inorganic polyphosphates production in Saccharomyces cerevisiae","authors":"Zipeng Chen ,&nbsp;Yanling Wu ,&nbsp;Lingfeng Qin ,&nbsp;Chen Wang ,&nbsp;Zhixin Li ,&nbsp;Xiaozhou Luo ,&nbsp;Wei Wei ,&nbsp;Jing Zhao","doi":"10.1016/j.synbio.2025.04.004","DOIUrl":"10.1016/j.synbio.2025.04.004","url":null,"abstract":"<div><div>Inorganic polyphosphate (polyP), a linear polymer of orthophosphate residues, plays critical roles in diverse biological processes spanning blood coagulation, immunomodulation, and post-translational protein modifications in eukaryotes. Notably, long-chain polyP (&gt;100 phosphate units) exhibits distinct biological functionalities compared to shorter-chain counterparts. While <em>Saccharomyces cerevisiae</em> serves as a promising microbial platform for polyP biosynthesis, the genetic regulatory mechanisms underlying polyP metabolism remain poorly elucidated. Here, we systematically investigated the genetic determinants governing intracellular polyP levels and chain length dynamics in yeast. Through screening a library of 55 single-gene knockout strains, we identified six mutants (<em>Δddp1</em>, <em>Δvip1</em>, <em>Δppn1</em>, <em>Δppn2</em>, <em>Δecm33</em>, and <em>Δccr4</em>) exhibiting elevated polyP accumulation, whereas deletions of <em>vtc1</em>, <em>kcs1</em>, <em>vma22</em>, <em>vma5</em>, <em>pho85</em>, <em>vtc4</em>, <em>vma2</em>, <em>vma3</em>, <em>ecm14</em>, and <em>vph2</em> resulted in near-complete polyP depletion. Subsequent combinatorial deletions in the <em>Δppn1</em> background revealed that the <em>Δppn1Δvip1</em> double mutant achieved synergistic enhancement in both polyP concentration (53.01 mg-P/g-DCW) and chain length, attributable to increased ATP availability and reduced polyphosphatase activity. Leveraging CRISPR/Cas9-mediated overexpression in <em>Δppn1Δvip1</em>, we engineered strain PP2 (<em>vtc4</em> overexpression), which demonstrated a 2-fold increase in polyP yield (62.6 mg-P/g-DCW) relative to wild-type BY4741, with predominant synthesis of long-chain species. Mechanistically, qRT-PCR analysis confirmed that PP2 exhibited 46-fold up-regulation of <em>vtc4</em> coupled with down-regulation of polyphosphatases encoding genes, <em>ppn2</em>, <em>ddp1</em>, and <em>ppx1</em>. This study performed a systematic study of regulating inorganic polyphosphates production in yeast and provides a synthetic biology strategy to engineer high-yield polyP-producing strains, advancing both fundamental understanding and biotechnological applications.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 816-826"},"PeriodicalIF":4.4,"publicationDate":"2025-04-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143848255","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":"Enhanced l-serine synthesis in Corynebacterium glutamicum by exporter engineering and Bayesian optimization of the medium composition","authors":"Yifan Huang ,&nbsp;Yujie Gao ,&nbsp;Yamin Huang ,&nbsp;Xiaogang Wang ,&nbsp;Meijuan Xu ,&nbsp;Guoqiang Xu ,&nbsp;Xiaojuan Zhang ,&nbsp;Hui Li ,&nbsp;Jinsong Shi ,&nbsp;Zhenghong Xu ,&nbsp;Xiaomei Zhang","doi":"10.1016/j.synbio.2025.04.003","DOIUrl":"10.1016/j.synbio.2025.04.003","url":null,"abstract":"<div><div><span>l</span>-serine is a versatile, high value-added amino acid, widely used in food, medicine and cosmetics. However, the low titer of <span>l</span>-serine has limited its industrial production. In this study, a cell factory without plasmid for efficient production of <span>l</span>-serine was constructed based on transport engineering. Firstly, the effects of <span>l</span>-serine exporter SerE overexpression and deletion on the cell growth and <span>l</span>-serine titer were investigated in <em>Corynebacterium glutamicum</em> (<em>C. glutamicum</em>) A36, overexpression of <em>s</em><em>erE</em> using a plasmid led to a 15.1% increase in <span>l</span>-serine titer but also caused a 15.1% decrease in cell growth. Subsequently, to increase the export capacity of SerE, we conducted semi-rational design and bioinformatics analysis, combined with alanine mutation and site-specific saturation mutation. The mutant E277K was obtained and exhibited a 53.2% higher export capacity compared to wild-type SerE, resulting in <span>l</span>-serine titer increased by 39.6%. Structural analysis and molecular dynamics simulations were performed to elucidate the mechanism. The results showed that the mutation shortened the hydrogen bond distance between the exporter and <span>l</span>-serine, enhanced complex stability, and reduced the binding energy. Finally, Bayesian optimization was employed to further improve <span>l</span>-serine titer of the mutant strain C-E277K. Under the optimized conditions, 47.77 g/L <span>l</span>-serine was achieved in a 5-L bioreactor, representing the highest reported titer for <em>C. glutamicum</em> to date. This study provides a basis for the transformation of <span>l</span>-serine export pathway and offers a new strategy for increasing <span>l</span>-serine titer.</div></div>","PeriodicalId":22148,"journal":{"name":"Synthetic and Systems Biotechnology","volume":"10 3","pages":"Pages 835-845"},"PeriodicalIF":4.4,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143850456","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}