Effective production of Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) by engineered Halomonas bluephagenesis grown on glucose and 1,4-Butanediol

Halomonas bluephagenesis has been engineered to produce flexible copolymers P34HB or poly(3-hydroxybutyrate-co-4-hydroxybutyrate) from glucose and petrol-chemical precursor, γ-butyrolactone. Herein, gene cluster aldD-dhaT was constructed in recombinant H. bluephagenesis for catalyzing 1,4-butanediol (BDO) into 4-hydroxybutyrate, which could grow to 86 g L⁻¹ dry cell mass (DCM) containing 77 wt% P(3HB-co-14 mol% 4HB) in 7-L bioreactor fed with glucose and bio-based BDO. Furthermore, 4HB monomer ratio could be increased to 16 mol% by engineered H. bluephagenesis TDH4-WZY254 with defected outer-membrane. Upon deletion of 4HB degradation pathway, followed by aldD-dhaT integration, the resulted H. bluephagenesis TDB141ΔAC was grown to 95 g L⁻¹ DCM containing 79 wt% P(3HB-co-14 mol% 4HB) with a BDO conversion efficiency of 86% under fed-batch fermentation. Notably, 4HB molar ratio can be significantly improved to 21 mol% with negligible effects on cell growth and P34HB synthesis by adding 50% more BDO. This study successfully demonstrated a fully bio-based P34HB effectively produced by H. bluephagenesis.