Combinatorial modulation of initial codons for improved zeaxanthin synthetic pathway efficiency in Escherichia coli

In this work, we found that by changing only the initiation codons, expression libraries of reporter proteins RFP, GFP, and lacZ, with a large dynamic range and evenly distributed expression levels, could be established inEscherichia coli. Subsequently, a novel combinatorial modulation of initial codon (CMIC) strategy was developed for metabolic pathway optimization. CMIC was applied to the zeaxanthin synthesis pathway inE.  coli, which achieved a nearly 10 ‐fold increase in zeaxanthin production. AbstractA balanced and optimized metabolic pathway is the basis for efficient production of a target metabolite. Traditional strategies mostly involve the manipulation of promoters or ribosome ‐binding sites, which can encompass long sequences and can be complex to operate. In this work, we found that by changing only the three nucleotides of the initiation codons, expression libraries of reporter proteins RFP, GFP, and lacZ with a large dynamic range and evenly distributed expression l evels could be established inEscherichia coli (E.  coli). Thus, a novel strategy that uses combinatorial modulation of initial codons (CMIC) was developed for metabolic pathway optimization and applied to the three genescrtZ, crtY, andcrtI of the zeaxanthin synthesis pathway inE.  coli. The initial codons of these genes were changed to random nucleotides NNN, and the gene cassettes were assembled into vectors via an optimized strategy based on type II restriction enzymes. With minimal labor ti...
Source: MicrobiologyOpen - Category: Microbiology Authors: Tags: ORIGINAL ARTICLE Source Type: research