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Adaptive laboratory evolution for improved tolerance of isobutyl acetate in Escherichia coli.
- Source :
-
Metabolic Engineering . Jan2022, Vol. 69, p50-58. 9p. - Publication Year :
- 2022
-
Abstract
- Previously, Escherichia coli was engineered to produce isobutyl acetate (IBA). Titers greater than the toxicity threshold (3 g/L) were achieved by using layer-assisted production. To avoid this costly and complex method, adaptive laboratory evolution (ALE) was applied to E. coli for improved IBA tolerance. Over 37 rounds of selective pressure, 22 IBA-tolerant mutants were isolated. Remarkably, these mutants not only tolerate high IBA concentrations, they also produce higher IBA titers. Using whole-genome sequencing followed by CRISPR/Cas9 mediated genome editing, the mutations (SNPs in metH , rho and deletion of arcA) that confer improved tolerance and higher titers were elucidated. The improved IBA titers in the evolved mutants were a result of an increased supply of acetyl-CoA and altered transcriptional machinery. Without the use of phase separation, a strain capable of 3.2-fold greater IBA production than the parent strain was constructed by combing select beneficial mutations. These results highlight the impact improved tolerance has on the production capability of a biosynthetic system. • E. coli was evolved for improved IBA tolerance, but also demonstrated higher titers. • Three mutations (metH , rho , arcA) were responsible for improved IBA tolerance. • Tolerance towards other acetate esters and acids was also demonstrated. • The metH , rho mutant had 3.2-fold greater production than the parent strain. [ABSTRACT FROM AUTHOR]
- Subjects :
- *ESCHERICHIA coli
*ACETATES
*NUCLEOTIDE sequencing
*PHASE separation
*TITERS
Subjects
Details
- Language :
- English
- ISSN :
- 10967176
- Volume :
- 69
- Database :
- Academic Search Index
- Journal :
- Metabolic Engineering
- Publication Type :
- Academic Journal
- Accession number :
- 154619556
- Full Text :
- https://doi.org/10.1016/j.ymben.2021.11.002