ACS Synthetic Biology最新文献_第2页

Redesigning CYP109E1 for Improving Catalytic Performance in 25-Hydroxyvitamin D₃ Synthesis Through Synergistic Enhancement of Electron Transfer and NADPH Regeneration.

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-04-04 DOI: 10.1021/acssynbio.4c00879

Jiaying Ai, Ziyang Yin, Jikai Gao, Wenjing Wang, Fuping Lu, Hui-Min Qin, Shuhong Mao

{"title":"Redesigning CYP109E1 for Improving Catalytic Performance in 25-Hydroxyvitamin D3 Synthesis Through Synergistic Enhancement of Electron Transfer and NADPH Regeneration.","authors":"Jiaying Ai, Ziyang Yin, Jikai Gao, Wenjing Wang, Fuping Lu, Hui-Min Qin, Shuhong Mao","doi":"10.1021/acssynbio.4c00879","DOIUrl":"https://doi.org/10.1021/acssynbio.4c00879","url":null,"abstract":"P450 enzymes are promising biocatalysts and play an important role in the field of drug synthesis due to their high catalytic activity and stereoselectivity. CYP109E1 from Bacillus megaterium was used to convert VD3 for the production of 25(OH)VD3. However, the industrial production was still limited due to the low catalytic performance of CYP109E1. To overcome this, we constructed an engineered strain containing a modified CYP109E1 coupled with an efficient electron transfer chain and NADPH regeneration system. First, Adx4-108T69E-Fpr was identified as the most compatible redox partner for the enzyme based on in-silico analysis. Then, targeted mutations were introduced at the substrate channel of CYP109E1, resulting in higher production efficiency. Next, the production of 25(OH)VD3 was increased by 13.1% after introducing a double Adx4-108T69E expression cassette. Finally, an NADPH regeneration system was introduced by overexpressing zwf, which increased the yield of 25(OH)VD3 48.7%. These results demonstrate that recombinant Escherichia coli BL21 (DE3) coexpressing CYP109E1_R70A-ZWF and 2Adx4-108T69Es-Fpr is an efficient whole-cell biocatalyst for the synthesis of 25(OH)VD3, illustrating an attractive strategy for improving the catalytic efficiency of P450 enzymes.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143778574","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}

引用次数: 0

Development of a Transposon-Based Genome Engineering Toolkit for Efficient and Adaptable Genetic Modifications in Wolfiporia cocos

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-04-02 DOI: 10.1021/acssynbio.4c0076610.1021/acssynbio.4c00766

Seungwoo Baek, Bogun Kim, Duleepa Pathiraja* and In-Geol Choi*,

{"title":"Development of a Transposon-Based Genome Engineering Toolkit for Efficient and Adaptable Genetic Modifications in Wolfiporia cocos","authors":"Seungwoo Baek, Bogun Kim, Duleepa Pathiraja* and In-Geol Choi*, ","doi":"10.1021/acssynbio.4c0076610.1021/acssynbio.4c00766","DOIUrl":"https://doi.org/10.1021/acssynbio.4c00766https://doi.org/10.1021/acssynbio.4c00766","url":null,"abstract":"Advances in genome engineering of fungal strains are rapidly progressing, driven by the increasing interest in fungal biotechnology. Given the unique genomic and cellular complexity of fungi, each strain benefits from a tailored toolkit for efficient genome engineering. Here, we present a transposon-based engineering toolkit specifically optimized for Wolfiporia cocos, a species valued for its bioactive compounds. This toolkit significantly improves transformation efficiency, enabling multiplexed gene integration and facilitating rapid, flexible prototyping by assembling multiple genes into transposomes in a cocktail format, which bypasses the need for an intricate genetic circuit assembly. Engineered strains demonstrated stable expression across generations, as confirmed by successful genomic integration. Additionally, we identified six native W. cocos promoters from transcriptomic data, with two showing robust, constitutive expression in the mycelium of engineered strains. This transposon-based toolkit offers a versatile resource for synthetic biology, supporting efficient and adaptable genetic modifications within fungal systems.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 4","pages":"1152–1160 1152–1160"},"PeriodicalIF":3.7,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143842331","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}

引用次数: 0

Marine Metagenome Mining Reveals Lanthipeptides Colwesin A-C, Exhibiting Novel Ring Topology and Anti-inflammatory Activity.

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-04-02 DOI: 10.1021/acssynbio.5c00057

Huimei Wang, Xing Zhao, Denghui Li, Liang Meng, Shanshan Liu, Youming Zhang, Liujie Huo

{"title":"Marine Metagenome Mining Reveals Lanthipeptides Colwesin A-C, Exhibiting Novel Ring Topology and Anti-inflammatory Activity.","authors":"Huimei Wang, Xing Zhao, Denghui Li, Liang Meng, Shanshan Liu, Youming Zhang, Liujie Huo","doi":"10.1021/acssynbio.5c00057","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00057","url":null,"abstract":"Marine natural products are promising sources for drug discovery due to their unique structures and diverse biological activities. The establishment of the Global Marine Microbiome Genome Catalogue (GOMC) has significantly expanded the repository of natural products derived from marine-associated bacteria. In this study, we identified the Class I lanthipeptide biosynthetic gene cluster col from Colwellia_A sp. based on the GOMC database. Through heterologous expression in Escherichia coli and subsequent structural analysis, we characterized three novel lanthipeptides, colwesins A-C, which possess unique cyclic structures characterized by an exceptionally large number of thioether rings. To the best of our knowledge, colwesin C is the first lanthipeptide simultaneously containing locked, nonoverlapped, and nested ring topologies. These findings highlight the robust ring-forming capacity of Class I lanthipeptide synthetases. Colwesins A-C were found to exhibit anti-inflammatory activity in lipopolysaccharide-induced mouse macrophage RAW264.7 cell lines without detectable cytotoxicity. Overall, our results broaden our understanding of the structural diversity of marine-derived lanthipeptides.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143762466","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}

引用次数: 0

Marine Metagenome Mining Reveals Lanthipeptides Colwesin A–C, Exhibiting Novel Ring Topology and Anti-inflammatory Activity

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-04-02 DOI: 10.1021/acssynbio.5c0005710.1021/acssynbio.5c00057

Huimei Wang, Xing Zhao, Denghui Li, Liang Meng, Shanshan Liu, Youming Zhang and Liujie Huo*,

{"title":"Marine Metagenome Mining Reveals Lanthipeptides Colwesin A–C, Exhibiting Novel Ring Topology and Anti-inflammatory Activity","authors":"Huimei Wang, Xing Zhao, Denghui Li, Liang Meng, Shanshan Liu, Youming Zhang and Liujie Huo*, ","doi":"10.1021/acssynbio.5c0005710.1021/acssynbio.5c00057","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00057https://doi.org/10.1021/acssynbio.5c00057","url":null,"abstract":"Marine natural products are promising sources for drug discovery due to their unique structures and diverse biological activities. The establishment of the Global Marine Microbiome Genome Catalogue (GOMC) has significantly expanded the repository of natural products derived from marine-associated bacteria. In this study, we identified the Class I lanthipeptide biosynthetic gene cluster col from Colwellia_A sp. based on the GOMC database. Through heterologous expression in Escherichia coli and subsequent structural analysis, we characterized three novel lanthipeptides, colwesins A–C, which possess unique cyclic structures characterized by an exceptionally large number of thioether rings. To the best of our knowledge, colwesin C is the first lanthipeptide simultaneously containing locked, nonoverlapped, and nested ring topologies. These findings highlight the robust ring-forming capacity of Class I lanthipeptide synthetases. Colwesins A–C were found to exhibit anti-inflammatory activity in lipopolysaccharide-induced mouse macrophage RAW264.7 cell lines without detectable cytotoxicity. Overall, our results broaden our understanding of the structural diversity of marine-derived lanthipeptides.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 4","pages":"1014–1020 1014–1020"},"PeriodicalIF":3.7,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143842270","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}

引用次数: 0

Development of a Transposon-Based Genome Engineering Toolkit for Efficient and Adaptable Genetic Modifications in Wolfiporia cocos.

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-04-02 DOI: 10.1021/acssynbio.4c00766

Seungwoo Baek, Bogun Kim, Duleepa Pathiraja, In-Geol Choi

{"title":"Development of a Transposon-Based Genome Engineering Toolkit for Efficient and Adaptable Genetic Modifications in Wolfiporia cocos.","authors":"Seungwoo Baek, Bogun Kim, Duleepa Pathiraja, In-Geol Choi","doi":"10.1021/acssynbio.4c00766","DOIUrl":"https://doi.org/10.1021/acssynbio.4c00766","url":null,"abstract":"Advances in genome engineering of fungal strains are rapidly progressing, driven by the increasing interest in fungal biotechnology. Given the unique genomic and cellular complexity of fungi, each strain benefits from a tailored toolkit for efficient genome engineering. Here, we present a transposon-based engineering toolkit specifically optimized for Wolfiporia cocos, a species valued for its bioactive compounds. This toolkit significantly improves transformation efficiency, enabling multiplexed gene integration and facilitating rapid, flexible prototyping by assembling multiple genes into transposomes in a cocktail format, which bypasses the need for an intricate genetic circuit assembly. Engineered strains demonstrated stable expression across generations, as confirmed by successful genomic integration. Additionally, we identified six native W. cocos promoters from transcriptomic data, with two showing robust, constitutive expression in the mycelium of engineered strains. This transposon-based toolkit offers a versatile resource for synthetic biology, supporting efficient and adaptable genetic modifications within fungal systems.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-04-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143770716","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}

引用次数: 0

A Staphylococcus aureus Virulence Inhibitor Identified by SaeRS Refactoring and Screening in Bacillus subtilis

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-04-01 DOI: 10.1021/acssynbio.4c0082610.1021/acssynbio.4c00826

Felix Ekness, Eric A. Wold, Catherine S. Leasure, Elena Musteata, Andrew J. Monteith, Clare Laut, Adriana E. Rosato, Eric P. Skaar and Jeffrey J. Tabor*,

{"title":"A Staphylococcus aureus Virulence Inhibitor Identified by SaeRS Refactoring and Screening in Bacillus subtilis","authors":"Felix Ekness, Eric A. Wold, Catherine S. Leasure, Elena Musteata, Andrew J. Monteith, Clare Laut, Adriana E. Rosato, Eric P. Skaar and Jeffrey J. Tabor*, ","doi":"10.1021/acssynbio.4c0082610.1021/acssynbio.4c00826","DOIUrl":"https://doi.org/10.1021/acssynbio.4c00826https://doi.org/10.1021/acssynbio.4c00826","url":null,"abstract":"Bacteria utilize two-component system (TCS) signal transduction pathways to sense environmental and physiological stimuli and mount appropriate responses. In opportunistic pathogens such as Staphylococcus aureus, TCSs activate virulence programs in response to host defense systems. Due to their critical role in pathogenesis, TCSs are important targets for antivirulence drug discovery campaigns. However, challenges associated with screening TCSs in pathogens and in vitro have limited the output of such efforts to a small number of characterized drug candidates. Here, we functionally express the S. aureus virulence-regulating TCS SaeRS from synthetic gene regulatory elements in the model bacterium Bacillus subtilis to reliably screen this system against a small molecule library under simple culturing conditions. Our approach reveals the compound NSC97920 as a strong inhibitor of SaeRS signaling. We combine in situ, in vivo, in silico, and in vitro characterization to demonstrate that NSC97920 suppresses the critical step of autophosphorylation in the SaeS histidine kinase, resulting in strong antivirulence activity. Our work shows that heterologous expression and screening of TCSs in model bacteria could accelerate the development of therapeutics against antibiotic-resistant pathogens.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 4","pages":"1191–1203 1191–1203"},"PeriodicalIF":3.7,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143842212","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}

引用次数: 0

A Staphylococcus aureus Virulence Inhibitor Identified by SaeRS Refactoring and Screening in Bacillus subtilis. 在枯草芽孢杆菌中通过 SaeRS 重构和筛选发现的金黄色葡萄球菌毒性抑制剂

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-04-01 DOI: 10.1021/acssynbio.4c00826

Felix Ekness, Eric A Wold, Catherine S Leasure, Elena Musteata, Andrew J Monteith, Clare Laut, Adriana E Rosato, Eric P Skaar, Jeffrey J Tabor

{"title":"A Staphylococcus aureus Virulence Inhibitor Identified by SaeRS Refactoring and Screening in Bacillus subtilis.","authors":"Felix Ekness, Eric A Wold, Catherine S Leasure, Elena Musteata, Andrew J Monteith, Clare Laut, Adriana E Rosato, Eric P Skaar, Jeffrey J Tabor","doi":"10.1021/acssynbio.4c00826","DOIUrl":"https://doi.org/10.1021/acssynbio.4c00826","url":null,"abstract":"Bacteria utilize two-component system (TCS) signal transduction pathways to sense environmental and physiological stimuli and mount appropriate responses. In opportunistic pathogens such as Staphylococcus aureus, TCSs activate virulence programs in response to host defense systems. Due to their critical role in pathogenesis, TCSs are important targets for antivirulence drug discovery campaigns. However, challenges associated with screening TCSs in pathogens and in vitro have limited the output of such efforts to a small number of characterized drug candidates. Here, we functionally express the S. aureus virulence-regulating TCS SaeRS from synthetic gene regulatory elements in the model bacterium Bacillus subtilis to reliably screen this system against a small molecule library under simple culturing conditions. Our approach reveals the compound NSC97920 as a strong inhibitor of SaeRS signaling. We combine in situ, in vivo, in silico, and in vitro characterization to demonstrate that NSC97920 suppresses the critical step of autophosphorylation in the SaeS histidine kinase, resulting in strong antivirulence activity. Our work shows that heterologous expression and screening of TCSs in model bacteria could accelerate the development of therapeutics against antibiotic-resistant pathogens.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143762464","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}

引用次数: 0

Dual Antibiotic-Free Plasmid Systems Enable High-Efficiency l-Fucose Biosynthesis.

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-03-27 DOI: 10.1021/acssynbio.5c00141

Jiawei Meng, Yingying Zhu, Zhen Lu, Wenli Zhang, Tao Zhang, Wanmeng Mu

{"title":"Dual Antibiotic-Free Plasmid Systems Enable High-Efficiency l-Fucose Biosynthesis.","authors":"Jiawei Meng, Yingying Zhu, Zhen Lu, Wenli Zhang, Tao Zhang, Wanmeng Mu","doi":"10.1021/acssynbio.5c00141","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00141","url":null,"abstract":"l-Fucose, a functional monosaccharide with significant commercial potential in the pharmaceutical, nutraceutical, and cosmetic industries, faces challenges in microbial production due to antibiotic-dependent plasmid maintenance systems. This study presents a dual antibiotic-free plasmid strategy in engineered Escherichia coli BL21(DE3) to achieve high-efficiency l-fucose biosynthesis. By integration of the hok/sok toxin-antitoxin system and a cysC-based auxotrophic selection into two plasmids, genetic stability and plasmid retention were ensured without antibiotics. Metabolic pathway optimization involved enhancing GDP-l-fucose supply via promoter replacements, genomic integration of key enzymes (α1,2-fucosyltransferase and α-l-fucosidase), and blocking l-fucose degradation. The engineered strain demonstrated robust performance, producing 7.99 g/L of l-fucose in shake-flask fermentation and 61.91 g/L via fed-batch cultivation─both antibiotic-free. This titer represents the highest reported l-fucose yield to date, highlighting the effectiveness of combining toxin-antitoxin and auxotrophic systems for sustainable, high-productivity microbial manufacturing.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":" ","pages":""},"PeriodicalIF":3.7,"publicationDate":"2025-03-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143727001","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}

引用次数: 0

Dual Antibiotic-Free Plasmid Systems Enable High-Efficiency l-Fucose Biosynthesis

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-03-27 DOI: 10.1021/acssynbio.5c0014110.1021/acssynbio.5c00141

Jiawei Meng, Yingying Zhu, Zhen Lu, Wenli Zhang, Tao Zhang and Wanmeng Mu*,

{"title":"Dual Antibiotic-Free Plasmid Systems Enable High-Efficiency l-Fucose Biosynthesis","authors":"Jiawei Meng, Yingying Zhu, Zhen Lu, Wenli Zhang, Tao Zhang and Wanmeng Mu*, ","doi":"10.1021/acssynbio.5c0014110.1021/acssynbio.5c00141","DOIUrl":"https://doi.org/10.1021/acssynbio.5c00141https://doi.org/10.1021/acssynbio.5c00141","url":null,"abstract":"l-Fucose, a functional monosaccharide with significant commercial potential in the pharmaceutical, nutraceutical, and cosmetic industries, faces challenges in microbial production due to antibiotic-dependent plasmid maintenance systems. This study presents a dual antibiotic-free plasmid strategy in engineered Escherichia coli BL21(DE3) to achieve high-efficiency l-fucose biosynthesis. By integration of the hok/sok toxin-antitoxin system and a cysC-based auxotrophic selection into two plasmids, genetic stability and plasmid retention were ensured without antibiotics. Metabolic pathway optimization involved enhancing GDP-l-fucose supply via promoter replacements, genomic integration of key enzymes (α1,2-fucosyltransferase and α-l-fucosidase), and blocking l-fucose degradation. The engineered strain demonstrated robust performance, producing 7.99 g/L of l-fucose in shake-flask fermentation and 61.91 g/L via fed-batch cultivation─both antibiotic-free. This titer represents the highest reported l-fucose yield to date, highlighting the effectiveness of combining toxin–antitoxin and auxotrophic systems for sustainable, high-productivity microbial manufacturing.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 4","pages":"1316–1322 1316–1322"},"PeriodicalIF":3.7,"publicationDate":"2025-03-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143842257","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}

引用次数: 0

Efficient De Novo Assembly of 100 kb-Scale Human Functional Immunoglobulin Heavy Variable (IGHV) Gene Fragments In Vitro.

IF 3.7 2区生物学

ACS Synthetic Biology Pub Date : 2025-03-26 DOI: 10.1021/acssynbio.5c00011

Haiqiong Li, Shuyao Xu, Yurui Liu, Yongqi Lu, Yunshan Ning

引用次数: 0