Advancing VB12 Production: Insights into Enhancing VB12 Titer in Ensifer adhaerens Casida A through ARTP Mutagenesis and Multiomics Analysis.

IF 3.7 2区 生物学 Q1 BIOCHEMICAL RESEARCH METHODS
Qi Wang, Yongheng Liu, Tengteng Zhu, Wei Zhao, Jianyu Su
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引用次数: 0

Abstract

Ensifer adhaerens, a microorganism recognized for its capacity to synthesize vitamin B12 (VB12), has garnered significant attention in recent years. Nonetheless, its practical application has been limited by low production yields. Atmospheric and room-temperature plasma (ARTP) mutagenesis was utilized to improve VB12 production and examine the associated mechanisms. Three high-yielding mutant strains─BCA-24, BCB-14, and BCC-27─were isolated through multiple rounds of mutagenesis. The VB12 titer of the highly productive mutant strain, BCA-24, rose significantly from 65.64 mg/L to 104.54 mg/L. Genome resequencing identified 14 mutated genes, of which seven (atpA, gntR, fusA, cobQ, ribD, cirA, and UP) were functionally validated through overexpression in wild-type strains and found to positively influence VB12 biosynthesis. Notably, coexpression of the cobQ and UP mutant genes in strain BCA-24 resulted in a VB12 titer of 163.68 mg/L. Transcriptomic analysis indicated that critical pathways related to energy metabolism, S-adenosylmethionine (SAM), 5-aminolevulinic acid (5-ALA), and riboflavin synthesis were significantly upregulated in BCA-24 relative to the wild type. A multiomics approach clarified the mechanisms through which these mutations increase VB12 production, including enhanced transcription and translation, optimized energy supply, and improved product efflux. The identification of novel candidate genes in Ensifer adhaerens, which have not been previously studied, provides valuable resources for future genetic engineering aimed at enhancing VB12 production efficiency. This study offers practical improvements in microbial VB12 production while also delivering essential insights into the genetic and metabolic regulation of this important biosynthetic pathway.

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来源期刊
CiteScore
8.00
自引率
10.60%
发文量
380
审稿时长
6-12 weeks
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.
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