Utilization of rare codon-rich markers for screening amino acid overproducers

• Published in: Nature communications Vol. 9; no. 1; pp. 3616 - 11
• Main Authors: Zheng, Bo, Ma, Xiaoyan, Wang, Ning, Ding, Tingting, Guo, Liwei, Zhang, Xiaorong, Yang, Yu, Li, Chun, Huo, Yi-Xin
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 06.09.2018; Nature Publishing Group; Nature Portfolio
• Subjects: 49; 49/47; 631/1647/2163; 631/326/2522; 631/326/41/2537; 631/61/318; 82/80; Amino acid starvation; Amino acids; Amino Acids - biosynthesis; Amino Acids - genetics; Antibiotic resistance; Antibiotics; Arginine; Codon; Codons; Corynebacterium glutamicum; Corynebacterium glutamicum - genetics; Corynebacterium glutamicum - growth & development; Corynebacterium glutamicum - metabolism; E coli; Escherichia coli - genetics; Escherichia coli - growth & development; Escherichia coli - metabolism; Frequency dependence; Genetic Engineering - methods; Genetic Markers; Green Fluorescent Proteins - genetics; Humanities and Social Sciences; L-Serine; Leucine; Markers; Microorganisms, Genetically-Modified; multidisciplinary; Proteins; Science; Science (multidisciplinary); Screening; Starvation; Translation; Translations
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-018-05830-0

The translation of rare codons relies on their corresponding rare tRNAs, which could not be fully charged under amino acid starvation. Theoretically, disrupted or retarded translation caused by the lack of charged rare tRNAs can be partially restored by feeding or intracellular synthesis of the corresponding amino acids. Inspired by this assumption, we develop a screening or selection system for obtaining overproducers of a target amino acid by replacing its common codons with the corresponding synonymous rare alternative in the coding sequence of selected reporter proteins or antibiotic-resistant markers. Results show that integration of rare codons can inhibit gene translations in a frequency-dependent manner. As a proof-of-concept, Escherichia coli strains overproducing l -leucine, l -arginine or l -serine are successfully selected from random mutation libraries. The system is also applied to Corynebacterium glutamicum to screen out l -arginine overproducers. This strategy sheds new light on obtaining and understanding amino acid overproduction strains. Current toxic analogues-based amino acid overproducer screening method cannot provide accuracy, sensitivity and high throughput simultaneously. Here, the authors use rare codon rich marker for the selection of overproducers of multiple amino acids from random mutation libraries of E. coli and C. glutamicum .