Biosensor-based multi-gene pathway optimization for enhancing the production of glycolate

Appl Environ Microbiol. 2021 Apr 9:AEM.00113-21. doi: 10.1128/AEM.00113-21. Online ahead of print.ABSTRACTGlycolate is widely used in industry, especially in the fields of chemical cleaning, cosmetics, and medical materials, and has broad market prospects for the future. Recent advances in metabolic engineering and synthetic biology have significantly improved the titer and yield of glycolate. However, an expensive inducer was used in previous studies that is not feasible for use in large-scale industrial fermentations. To constitutively biosynthesize glycolate, the expression level of each gene of the glycolate synthetic pathway needs to be systemically optimized. The main challenge of multi-gene pathway optimization is being able to select or screen the optimum strain from the randomly assembled library by an efficient high-throughput method within a short period of time. To overcome these challenges, we firstly established a glycolate-responsive biosensor and developed agar plate- and 48-well deep well plate-scale high-throughput screening methods for rapid screening of superior glycolate producers from a large library. A total of 22 gradient strength promoter-5'-UTR complexes were randomly cloned upstream of the genes of the glycolate synthetic pathway, generating a large random assembled library. After rounds of screening, the optimum strain was obtained from 6×105 transformants in a week, and it achieved a titer of 40.9 ± 3.7 g/L glycolate in a 5-L bioreactor. Further...
Source: Applied and Environmental Microbiology - Category: Microbiology Authors: Source Type: research