{"title":"Rational and Semirational Approaches for Engineering Salicylate Production in <i>Escherichia coli</i>.","authors":"Chenghu Chen, Cong Gao, Guipeng Hu, Wanqing Wei, Xiaoge Wang, Jian Wen, Xiulai Chen, Liming Liu, Wei Song, Jing Wu","doi":"10.1021/acssynbio.4c00366","DOIUrl":null,"url":null,"abstract":"<p><p>Salicylate plays a pivotal role as a pharmaceutical intermediate in drugs, such as aspirin and lamivudine. The low catalytic efficiency of key enzymes and the inherent toxicity of salicylates to cells pose significant challenges to large-scale microbial production. In this study, we introduced the salicylate synthase Irp9 into an l-phenylalanine-producing <i>Escherichia coli</i>, constructing the shortest salicylate biosynthetic pathway. Subsequent protein engineering increased the catalytic efficiency of Irp9 by 33.5%. Furthermore, by integrating adaptive evolution with transcriptome analysis, we elucidated the crucial mechanism of efflux proteins in salicylate tolerance. The elucidation of this mechanism guided us in the targeted modification of these transport proteins, achieving a reported maximum level of 3.72 g/L of salicylate in a shake flask. This study highlights the importance of efflux proteins for enhancing the productivity of microbial cell factories in salicylate production, which also holds potential for application in the green synthesis of other phenolic acids.</p>","PeriodicalId":26,"journal":{"name":"ACS Synthetic Biology","volume":null,"pages":null},"PeriodicalIF":3.7000,"publicationDate":"2024-10-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"ACS Synthetic Biology","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1021/acssynbio.4c00366","RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 0
Abstract
Salicylate plays a pivotal role as a pharmaceutical intermediate in drugs, such as aspirin and lamivudine. The low catalytic efficiency of key enzymes and the inherent toxicity of salicylates to cells pose significant challenges to large-scale microbial production. In this study, we introduced the salicylate synthase Irp9 into an l-phenylalanine-producing Escherichia coli, constructing the shortest salicylate biosynthetic pathway. Subsequent protein engineering increased the catalytic efficiency of Irp9 by 33.5%. Furthermore, by integrating adaptive evolution with transcriptome analysis, we elucidated the crucial mechanism of efflux proteins in salicylate tolerance. The elucidation of this mechanism guided us in the targeted modification of these transport proteins, achieving a reported maximum level of 3.72 g/L of salicylate in a shake flask. This study highlights the importance of efflux proteins for enhancing the productivity of microbial cell factories in salicylate production, which also holds potential for application in the green synthesis of other phenolic acids.
期刊介绍:
The journal is particularly interested in studies on the design and synthesis of new genetic circuits and gene products; computational methods in the design of systems; and integrative applied approaches to understanding disease and metabolism.
Topics may include, but are not limited to:
Design and optimization of genetic systems
Genetic circuit design and their principles for their organization into programs
Computational methods to aid the design of genetic systems
Experimental methods to quantify genetic parts, circuits, and metabolic fluxes
Genetic parts libraries: their creation, analysis, and ontological representation
Protein engineering including computational design
Metabolic engineering and cellular manufacturing, including biomass conversion
Natural product access, engineering, and production
Creative and innovative applications of cellular programming
Medical applications, tissue engineering, and the programming of therapeutic cells
Minimal cell design and construction
Genomics and genome replacement strategies
Viral engineering
Automated and robotic assembly platforms for synthetic biology
DNA synthesis methodologies
Metagenomics and synthetic metagenomic analysis
Bioinformatics applied to gene discovery, chemoinformatics, and pathway construction
Gene optimization
Methods for genome-scale measurements of transcription and metabolomics
Systems biology and methods to integrate multiple data sources
in vitro and cell-free synthetic biology and molecular programming
Nucleic acid engineering.