A microbial factory for lactate-based polyesters using a lactate-polymerizing enzyme

TLDR

Polylactate (PLA) is a representative bio‑based polyester produced by chemo‑bio processes that polymerize lactate from microbial fermentation, and an engineered PHA synthase was identified in a two‑phase in‑vitro polymerization system as a potential catalyst. The study aims to develop a one‑step microbial route to lactate‑based polyesters by testing whether PHA synthase can polymerize lactate‑CoA. An engineered *E. coli* strain producing LA‑CoA via a CoA transferase was combined with a modified PHA synthase, enabling one‑step biosynthesis of a lactate‑containing copolyester (P(6 mol % LA‑co‑94 mol % 3HB)) with a number‑average molecular weight of 1.9 × 10⁵, verified by GPC, GC/MS, and NMR.

Abstract

Polylactate (PLA) is synthesized as a representative bio-based polyester by the chemo-bio process on the basis of metal catalyst-mediated chemical polymerization of lactate (LA) supplied by microbial fermentation. To establish the one-step microbial process for synthesis of LA-based polyesters, we explored whether polyhydroxyalkanoate (PHA) synthase would exhibit polymerizing activity toward a LA-coenzyme A (CoA), based on the fact that PHA monomeric constituents, especially 3-hydroxybutyrate (3HB), are structurally analogous to LA. An engineered PHA synthase was discovered as a candidate by a two-phase in vitro polymerization system previously developed. An LA-CoA producing Escherichia coli strain with a CoA transferase gene was constructed, and the generation of LA-CoA was demonstrated by capillary electrophoresis/MS analysis. Next, when the engineered PHA synthase gene was introduced into the resultant recombinant strain, we confirmed the one-step biosynthesis of the LA-incorporated copolyester, P(6 mol% LA-co-94 mol% 3HB), with a number-average molecular weight of 1.9 x 10(5), as revealed by gel permeation chromatography, gas chromatography/MS, and NMR.