(571g) Renewable Production of 5-Carbon Polyamide Building Blocks Using Engineered E. Coli

Polyamides are a class of homopolymers and copolymers with properties that balance excellent mechanical strength with durability and chemical resistance. Common examples include nylons which are produced from amino acid or diamine and diacid monomer building blocks. Today, such monomers are principally derived from petroleum resources.  In recent years, however, the microbial production of 4- through 6-carbon monomers such as succinic acid, adipic acid, putrescine, and cadaverine have been reported. Here we report how, through de novo pathway engineering, our lab has been exploring the ‘bottom up’ development of microbial biocatalysts to produce other remaining polyamide monomer building blocks from renewable feedstocks.  Systematic over-expression of the natural lysine degradation pathway enzymes of Pseudomonas sp. in a lysine over-producing Escherichia coli strain has allowed us to produce, as individual products, both 5-aminovaleric acid and glutaric acid (precursors to nylon-5 and copolymers such as nylon-5,5, for example).  To date, genes from the davBADT operon of P. putida KT2440 has proven to be support the highest productivities in shake flask cultures.  Alternative expression strategies been designed to compare the utility of the natural operon versus the individualized expression of pathway enzymes with the latter proving most effective.  In shake flask cultures containing glucose in minimal media, our initial strains have already achieved 5-aminovaleric acid and glutaric acid titers as high as at least 350 mg/L. Here we report an overview of our most recent findings in this project. In all cases of strain development, systematic pathway development will be described through the recruitment and screening of candidate isoenzymes from different genetic sources for each step.  In addition, to support greater overall productivity, strategies aimed at increasing precursor availability, improving pathway flux, and overcoming product toxicity are also described.