ACS Synthetic Biology最新文献_第7页

Directed Evolution of Branched-Chain α-Keto Acid Decarboxylase for 3-Hydroxypropionic Acid Production in Escherichia coli via Oxaloacetate 大肠杆菌经草酰乙酸生产3-羟基丙酸支链α-酮酸脱羧酶的定向进化

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-15 DOI: 10.1021/acssynbio.5c00267

Chuang Wang, René C. L. Olsthoorn and Huub J. M. de Groot*,

{"title":"Directed Evolution of Branched-Chain α-Keto Acid Decarboxylase for 3-Hydroxypropionic Acid Production in Escherichia coli via Oxaloacetate","authors":"Chuang Wang, René C. L. Olsthoorn and Huub J. M. de Groot*, ","doi":"10.1021/acssynbio.5c00267","DOIUrl":"10.1021/acssynbio.5c00267","url":null,"abstract":"3-Hydroxypropionic acid (3-HP) serves as a crucial platform chemical with diverse applications across various industries. In this study, the oxaloacetate pathway was utilized for 3-HP production. This pathway involves the decarboxylation of oxaloacetate into malonic semialdehyde, catalyzed by branched-chain α-keto acid decarboxylase (KdcA), which is subsequently reduced to 3-HP by dehydrogenases. Directed evolution of KdcA was carried out to enhance its catalytic efficiency toward oxaloacetate, resulting in a KdcAM8 mutant with the following substitutions: S286R, S287T, F381H, F382P, L534S, L535F, M538T, and G539F. Compared to wild-type (WT) KdcA, KdcAM8 exhibits a lower KM value toward oxaloacetate (KM = 1.15 mM vs KM > 25 mM). Among these mutations, the single mutants S286R and S287T exhibited 5.5-fold and 1.3-fold increased activities, respectively. Instead of WT KdcA, the KdcAM8 mutant was integrated into Escherichia coli (E. coli) BL21 strain, resulting in the production of 3-HP at a concentration of 0.11 mM. To further improve 3-HP production, two dehydrogenases were compared for the downstream conversion of malonic semialdehyde into 3-HP, and two carboxylases were explored to enhance the upstream precursor supply of oxaloacetate. Additionally, the growth conditions were optimized. Finally, a nonnatural oxaloacetate pathway was successfully engineered in the E. coli BL21 strain, achieving a 3-HP titer of approximately 0.71 mM from glucose. This work illustrates that protein engineering is a powerful tool for modulating flux in the target pathway and holds promise for the future development of the oxaloacetate pathway to improve the 3-HP yield.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3487–3496"},"PeriodicalIF":3.9,"publicationDate":"2025-08-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.acs.org/doi/pdf/10.1021/acssynbio.5c00267","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144853945","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}

引用次数: 0

Multiplexed Genome Editing and Transcriptional Knockdown in Yarrowia lipolytica by CRISPR-Cpf1 and an Orthogonal T7 System 利用CRISPR-Cpf1和正交T7系统多重基因组编辑和转录敲低脂耶氏菌。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-14 DOI: 10.1021/acssynbio.5c00104

Hanqing Zhang, Luai R. Khoury and Peng Xu*,

{"title":"Multiplexed Genome Editing and Transcriptional Knockdown in Yarrowia lipolytica by CRISPR-Cpf1 and an Orthogonal T7 System","authors":"Hanqing Zhang, Luai R. Khoury and Peng Xu*, ","doi":"10.1021/acssynbio.5c00104","DOIUrl":"10.1021/acssynbio.5c00104","url":null,"abstract":"Yarrowia lipolytica, a nonconventional yeast, has become an industrial workhorse to synthesize valuable compounds, including lipids, oleochemicals, and nutraceuticals. While the synthetic biology toolkits to engineer the genome and endogenous metabolic pathways are not as developed as Baker’s yeast, it has emerged as a promising microbial host for industrial applications. In this study, we examined the multiplexed editing capability of the CRISPR-AsCpf1 coupled with gRNAs generated from either a yeast native promoter or an orthogonal T7 promoter, which yielded 73.3% editing efficiency for up to four target genes and 100% editing efficiency for two genes. We also attempted two strategies to enhance homology-directed recombination (HDR) efficiency; only minor improvements were observed. We further demonstrated that CRISPR-dAsCpf1 with T7-driven gRNA achieved significant gene knockdown compared to the CRISPR-RfxCas13d system. Its knockdown efficiency was comparable to that of an antisense T7 promoter system. Taken together, this work provides a facile toolkit that enables efficient and multiplexed genome editing and transcriptional knockdown of critical genes by combining CRISPR-Cpf1 with an orthogonal T7 transcription system in Y. lipolytica.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3377–3386"},"PeriodicalIF":3.9,"publicationDate":"2025-08-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144853946","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 High-Sensitivity Genetically Encoded Biosensor for Terephthalic Acid Detection in PET Degradation 用于PET降解对苯二甲酸检测的高灵敏度遗传编码生物传感器。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-14 DOI: 10.1021/acssynbio.5c00279

Seok Jin Oh, Jung-Ung An, Jun-Hong Park, Eun-Woo Choi, Seong Keun Kim, Seung Gyun Woo, Tae Hyun Kim, Bong Hyun Sung, Seung-Goo Lee, Kil Koang Kwon* and Dae-Hee Lee*,

{"title":"A High-Sensitivity Genetically Encoded Biosensor for Terephthalic Acid Detection in PET Degradation","authors":"Seok Jin Oh, Jung-Ung An, Jun-Hong Park, Eun-Woo Choi, Seong Keun Kim, Seung Gyun Woo, Tae Hyun Kim, Bong Hyun Sung, Seung-Goo Lee, Kil Koang Kwon* and Dae-Hee Lee*, ","doi":"10.1021/acssynbio.5c00279","DOIUrl":"10.1021/acssynbio.5c00279","url":null,"abstract":"The accumulation of polyethylene terephthalate (PET) waste poses a serious environmental challenge due to its durability and resistance to degradation. Enzymatic PET hydrolysis offers a sustainable solution, but efficient high-throughput screening tools for PET-degrading enzymes remain limited. Here, we report a genetically encoded biosensor (GEB) for terephthalic acid (TPA)─the primary monomer released during PET degradation─that enables rapid and sensitive detection of enzymatic activity. We engineered a TphR-based biosensor in Escherichia coli, combining an optimized transcriptional system with diverse TPA uptake transporters to enhance intracellular TPA accumulation. This dual strategy improved the signal intensity and broadened the detection range. The best-performing configuration, integrating a high-affinity transporter with fine-tuned genetic components, achieved a detection limit of 1  μM TPA─a 1,000-fold sensitivity improvement over the initial design. We validated the system using PETases, including Ideonella sakaiensis-derived FAST-PETase, and benchmarked it against HPLC assays. The biosensor reliably distinguished PETase variants based on hydrolytic activity, demonstrating its utility for directed evolution, metagenomic screening, and enzyme engineering. This work establishes a rapid, scalable, and ultrasensitive biosensor platform for monitoring PET hydrolysis. The engineered GEB offers a robust, low-cost alternative to conventional analytics, accelerating the discovery and optimization of PET-degrading enzymes for plastic upcycling and circular bioeconomy applications.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3497–3509"},"PeriodicalIF":3.9,"publicationDate":"2025-08-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144853943","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

Unraveling the Constrained Cell Growth in Engineered Living Materials 揭示工程生物材料中受约束的细胞生长。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-14 DOI: 10.1021/acssynbio.5c00378

Shuang-Shuang Sun, Cheng-Cheng Ding, Hai-Yan Yu, Xian-Zheng Yuan, Shu-Guang Wang* and Peng-Fei Xia*,

{"title":"Unraveling the Constrained Cell Growth in Engineered Living Materials","authors":"Shuang-Shuang Sun, Cheng-Cheng Ding, Hai-Yan Yu, Xian-Zheng Yuan, Shu-Guang Wang* and Peng-Fei Xia*, ","doi":"10.1021/acssynbio.5c00378","DOIUrl":"10.1021/acssynbio.5c00378","url":null,"abstract":"Engineered living materials (ELMs) leverage the integrative advantages of materials science and synthetic biology for advanced functionalities. Predicting and controlling cellular behavior are essential for designing and building ELMs, requiring a fundamental understanding of the growth dynamics of encapsulated cells. Here, we interrogate the interference of constrained growth with the engineered functionalities and cellular physiology of cyanobacteria and unveil the dynamic interaction between cell growth and spatial confinements within photosynthetic ELMs. We observed that engineered cyanobacteria within ELMs exhibited compromised performances in growth, uptake of nonutilizable substrate, and synthesis of customized products, while ELMs could protect encapsulated cells from external stresses. Besides commonly accepted external influences, we identified abnormally high levels of reactive oxygen species and impaired oxygen photosynthesis inside the cells encapsulated in the ELMs. Finally, we illustrated the dynamics of cell growth within the confined spaces enveloped by the material matrices, forming clustered cell aggregates and compressed growth bubbles until the spatial limits. Our study provides a fundamental yet often overlooked connection between cellular behavior and spatial confinement, consolidating the foundation for advanced ELM innovations.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3600–3611"},"PeriodicalIF":3.9,"publicationDate":"2025-08-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144843792","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

Bioengineering of Probiotic Yeast Saccharomyces boulardii for Advanced Biotherapeutics 先进生物治疗中益生菌酵母的生物工程研究。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-14 DOI: 10.1021/acssynbio.5c00236

Tiew-Yik Ting, Wei-Jing Lee and Hoe-Han Goh*,

{"title":"Bioengineering of Probiotic Yeast Saccharomyces boulardii for Advanced Biotherapeutics","authors":"Tiew-Yik Ting, Wei-Jing Lee and Hoe-Han Goh*, ","doi":"10.1021/acssynbio.5c00236","DOIUrl":"10.1021/acssynbio.5c00236","url":null,"abstract":"Saccharomyces cerevisiae var. boulardii (Sb), a subspecies of S. cerevisiae (Sc), is widely recognized for its probiotic properties. Recently, Sb has attracted growing interest as a chassis organism for engineered live biotherapeutics and advanced microbiome therapies. Traditional genetic manipulation techniques developed for Sc are now being successfully adapted for Sb, facilitating diverse genome integration strategies to enable the in situ biomanufacturing of functional molecules for disease intervention. Concurrently, research efforts are advancing Sb’s potential as a platform for biosensing applications and diagnostic tools through the development of disease-responsive biosensors. Biosafety concerns are also being addressed through the design of biocontainment strains that ensure controlled application. To the best of our knowledge, earlier reviews have largely emphasized its clinical applications, safety profile, and probiotic mechanisms. This review uniquely consolidates recent advances in genetic modification, metabolic engineering, and synthetic biology strategies applied to Sb for therapeutic use. Together, these synthetic biology advancements position Sb as a promising and versatile platform for next-generation microbiome-based therapeutics and expanding applications in human health and food biotechnology.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3275–3292"},"PeriodicalIF":3.9,"publicationDate":"2025-08-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144853944","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

Transcriptomics-Driven Engineering of Yarrowia lipolytica for Enhanced Fatty Acid Biosynthesis 转录组学驱动的多脂耶洛氏菌促进脂肪酸生物合成工程。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-14 DOI: 10.1021/acssynbio.5c00307

He Qiao, Zaihang Dong, Bofan Yu, Xiao Hong* and Xuye Lang*,

{"title":"Transcriptomics-Driven Engineering of Yarrowia lipolytica for Enhanced Fatty Acid Biosynthesis","authors":"He Qiao, Zaihang Dong, Bofan Yu, Xiao Hong* and Xuye Lang*, ","doi":"10.1021/acssynbio.5c00307","DOIUrl":"10.1021/acssynbio.5c00307","url":null,"abstract":"While Yarrowia lipolytica has gained prominence as a microbial chassis for biomanufacturing, its broader application faces two critical limitations: incomplete genetic annotation and insufficient characterization of regulatory elements, rendering the construction of high-efficiency microbial cell factories a time-consuming and empirically driven process. Notably, vast transcriptomic data sets in public database remain underutilized for systematic gene discovery. To address these limitations, we developed Findgene─a computational pipeline integrating standardized transcriptomic meta-analysis with weighted gene coexpression network analysis (WGCNA). Application of this tool to consolidated Y. lipolytica data sets identified six candidate regulatory genes related to fatty acid metabolism: YALI0B12342g, YALI0A07733g, YALI0C03003g, YALI0C16797g, YALI0A20207g, and YALI0D01001g. Remarkably, substitution YALI0B12342g with the G643R mutant increased total fatty acid production by 131%. Meanwhile, experimental validation revealed that plasmid-mediated overexpression of YALI0A07733g and YALI0A20207g significantly enhanced total fatty acid titer. Based on these, the combinatorial engineering strategy incorporating overexpression of YALI0A07733g/YALI0A20207g and implementation of the YALI0B12342g G643R variant achieved a 2.9-fold enhancement in total fatty acid production compared to wild type Po1f strains. This optimized chassis demonstrates substantial potential for scale-up production of fatty acid-derived compounds. Furthermore, the FindGene framework establishes a generalized methodology for regulatory gene efficient and economical discovery that could be adapted to engineer other nonconventional yeast species.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3520–3529"},"PeriodicalIF":3.9,"publicationDate":"2025-08-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144853947","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

Genetic Memory Devices to Detect Specialized Metabolites in Plant and Soil Microbiomes 检测植物和土壤微生物组中特殊代谢物的遗传记忆装置。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-13 DOI: 10.1021/acssynbio.5c00073

Morten Lindqvist Hansen, Kristoffer Ioannis Tang Kordatos, Jens-Jakob Krogh Nørgaard, Johan Peter Bredal Jørgensen, Mikael Lenz Strube, Morten Dencker Schostag, Lei Yang and Lars Jelsbak*,

{"title":"Genetic Memory Devices to Detect Specialized Metabolites in Plant and Soil Microbiomes","authors":"Morten Lindqvist Hansen, Kristoffer Ioannis Tang Kordatos, Jens-Jakob Krogh Nørgaard, Johan Peter Bredal Jørgensen, Mikael Lenz Strube, Morten Dencker Schostag, Lei Yang and Lars Jelsbak*, ","doi":"10.1021/acssynbio.5c00073","DOIUrl":"10.1021/acssynbio.5c00073","url":null,"abstract":"Root-associated microbiomes significantly influence plant growth and resilience through intricate chemical dialogues mediated by plant- and microbe-derived specialized metabolites. These metabolites play pivotal roles in shaping the assembly, dynamics, and ecological functions of soil microbiomes. Despite advances in in vitro and DNA sequencing studies, a comprehensive understanding of in situ chemical signaling within plant and soil microbiomes remains elusive due to experimental constraints. To address this gap, we developed and tuned a set of five whole-cell biosensors in Escherichia coli for spatiotemporal, nondisruptive detection of biologically relevant specialized metabolites, including 2,4-diacetylphloroglucinol, pyoluteorin, tetracycline, salicylic acid, and naringenin. Four of these biosensors were successfully adapted to the soil-compatible Pseudomonas putida KT2440 Δall-Φ strain. Additionally, the four sensors were shown to respond to their cognate ligand in a nonsterile soil extract medium containing a diverse microbiome extracted from soil. By employing genetic memory devices with DNA barcodes for readouts, our approach provides a scalable platform for sensing additional specialized metabolites in the future. This work demonstrates the potential of biosensor technologies to unravel the complex chemical interactions driving soil microbiome ecology, with implications for sustainable agricultural practices.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3362–3376"},"PeriodicalIF":3.9,"publicationDate":"2025-08-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144843790","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

Innovations in Yeast Synthetic Biology: Engineered Discovery Systems for Immunotherapy 酵母合成生物学的创新：免疫治疗的工程发现系统。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-13 DOI: 10.1021/acssynbio.5c00321

Ethan W. Slaton, Natalie Clay, Nathan Phan and Blaise R. Kimmel*,

{"title":"Innovations in Yeast Synthetic Biology: Engineered Discovery Systems for Immunotherapy","authors":"Ethan W. Slaton, Natalie Clay, Nathan Phan and Blaise R. Kimmel*, ","doi":"10.1021/acssynbio.5c00321","DOIUrl":"10.1021/acssynbio.5c00321","url":null,"abstract":"Yeast-based platforms are emerging as innovative synthetic biology tools for the discovery of immunotherapeutic proteins. Through the integration of (i) high-throughput surface display technologies, (ii) automated evolution systems (such as OrthoRep), and (iii) computational design strategies, the field of synthetic biology can make a direct impact toward rapidly identifying and engineering novel protein-based therapeutics. In this review, we will highlight the latest innovations regarding using engineered yeast to display proteins (e.g., nanobodies) and screen for potential antigens for immune receptors (e.g., GPCRs, TCRs). We will also discuss emerging areas in which the field has recently progressed and how the innovative technologies from these efforts help bridge the gap between synthetic biology and immunology such as identifying therapeutic binding events for engineered proteins of interest with the potential to actuate downstream immune responses. These innovations illustrate how yeast enables new design, build, test, and learn (DBTL) workflows in immunoengineering and offers a scalable, programmable chassis for developing tools and technologies for the construction of next-generation biotherapeutics.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3293–3305"},"PeriodicalIF":3.9,"publicationDate":"2025-08-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144843791","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

Cell-Free Production of Soybean Leghemoglobins and Nonsymbiotic Hemoglobin 大豆血红蛋白和非共生血红蛋白的无细胞生产。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-12 DOI: 10.1021/acssynbio.5c00197

Amanda P. Rocha, Mariele A. Palmeiras, Marco Antônio deOliveira, Lilian H. Florentino, Thais R. Cataldi, Daniela M. de C. Bittencourt, Carlos A. Labate, Gracia M. S. Rosinha and Elibio L. Rech*,

{"title":"Cell-Free Production of Soybean Leghemoglobins and Nonsymbiotic Hemoglobin","authors":"Amanda P. Rocha, Mariele A. Palmeiras, Marco Antônio deOliveira, Lilian H. Florentino, Thais R. Cataldi, Daniela M. de C. Bittencourt, Carlos A. Labate, Gracia M. S. Rosinha and Elibio L. Rech*, ","doi":"10.1021/acssynbio.5c00197","DOIUrl":"10.1021/acssynbio.5c00197","url":null,"abstract":"Hemoglobins are heme proteins and are present in certain microorganisms, higher plants, and mammals. Two types of hemoglobin are found in legume nodules: leghemoglobin (LegH) or symbiotic and nonsymbiotic (nsHb). LegHs occur in high amounts in legume roots, and together with bacteroides, are responsible for the nitrogen fixation process. nsHb Class 1 proteins have very high affinity for O2 and are found in monocotyledons and legumes. LegH has attracted great interest in the vegetable meat industry owing to its organoleptic and nutritional properties. In this study, soybean LegHs A, C1, C2 and C3 and nsHb were produced via Escherichia coli-based cell-free systems (CFS) and their amino acid sequences were correctly synthesized. In addition, certain post-translational modifications were made, which were confirmed using liquid chromatography–mass spectrometry analysis. All LegHs produced in this system exhibited peroxidase activity and heme binding, which were correlated with their concentrations in the assays. Furthermore, all proteins were readily digested by pepsin within 1 min under analog digestion conditions. Thus, LegHs and nsHb proteins were produced in this study using cell-free systems, maintaining their functionality and digestibility. These findings suggest that they could serve as viable alternative food additives for plant-based meat.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3445–3456"},"PeriodicalIF":3.9,"publicationDate":"2025-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.acs.org/doi/pdf/10.1021/acssynbio.5c00197","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144820115","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}

引用次数: 0

Functionalizing Nisin with a Sugar Moiety Improves Its Solubility and Results in an Altered Antibacterial Spectrum and Mode of Action 用糖片段功能化Nisin提高其溶解度，并导致抗菌光谱和作用方式的改变。

IF 3.9 2区生物学

ACS Synthetic Biology Pub Date : 2025-08-12 DOI: 10.1021/acssynbio.5c00353

Longcheng Guo, Oscar P. Kuipers and Jaap Broos*,

{"title":"Functionalizing Nisin with a Sugar Moiety Improves Its Solubility and Results in an Altered Antibacterial Spectrum and Mode of Action","authors":"Longcheng Guo, Oscar P. Kuipers and Jaap Broos*, ","doi":"10.1021/acssynbio.5c00353","DOIUrl":"10.1021/acssynbio.5c00353","url":null,"abstract":"Glycosylation, a widespread post-translational modification, is present in all kingdoms of life. Despite the extensive structural diversity found in ribosomally synthesized and post-translationally modified peptides (RiPPs), only a few glycosylated bacteriocins, known as glycocins, have been identified. Notably, glycocins such as glycocin F, ASM1, and enterocin F4-9, exhibit antimicrobial properties and distinct glycoactivity, indicating that glycosylation is crucial for their bioactivity. The development of practical, and widely applicable systems for glycosylation of RiPPs is therefore highly desirable. In this study, we introduce an expression system that utilizes Lactococcus lactis as a host for the efficient incorporation of the noncanonical amino acid homopropargylglycine (Hpg) into the well-studied RiPP nisin, and some structurally related variants. Hpg, which has an alkyne functional group, allows for further chemical modifications with azido-sugar containing substrates through click chemistry. We reveal that glycosylated nisin at position 17 shows strong activity against Enterococcus faecium strains, but its activity against other pathogens such as Staphylococcus aureus, Enterococcus faecalis, and Bacillus cereus is reduced. Moreover, mode of action studies show that the addition of sugar diminishes its typical pore-forming ability of nisin against E. faecium while preserving its lipid II binding ability. Interestingly, the addition of a hydrophilic sugar significantly enhances its water solubility around 4-fold at neutral pH, indicating potential for improved drug applications. These findings highlight the potential of this methodology for glycosylation of RiPPs, leading to the creation of new antimicrobial products with varied characteristics. This also broadens the toolkit for enhancing and discovering peptide-based drugs.","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":"14 9","pages":"3568–3577"},"PeriodicalIF":3.9,"publicationDate":"2025-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.acs.org/doi/pdf/10.1021/acssynbio.5c00353","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144820116","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}

引用次数: 0