Biosynthesis of P(3HB-co-3HHx) Copolymers by a Newly Engineered Strain of Cupriavidus necator PHB−4/pBBR_CnPro-phaCRp for Skin Tissue Engineering Application

Abstract

Polyhydroxyalkanoates (PHAs) are biodegradable polymers synthesized by certain bacteria and archaea with functions comparable to conventional plastics. Previously, our research group reported a newly PHA-producing bacterial strain, <i>Rhodococcus pyridinivorans</i> BSRT1-1, from the soil in Thailand. However, this strain's PHA synthase (<i>phaC</i><i>_Rp</i>) gene has not yet been characterized. Thus, this study aims to synthesize PHA using a newly engineered bacterial strain, <i>Cupriavidus necator</i> PHB^-4/pBBR_CnPro-<i>phaC_Rp</i>, which harbors the <i>phaC</i><i>_Rp</i> from strain BSRT1-1, and characterize the properties of PHA for skin tissue engineering application. To the best of our knowledge, this is the first study on the characterization of the PhaC from <i>R. pyridinivorans</i> species. The results demonstrated that the expression of the <i>phaC</i><i>_Rp</i> in <i>C. necator</i> PHB^-4 had developed in PHA production up to 3.1 ± 0.3 g/L when using 10 g/L of crude palm kernel oil (CPKO) as a sole carbon source. Interestingly, the engineered strain produced a 3-hydroxybutyrate (3HB) with 2 mol% of 3-hydroxyhexanoate (3HHx) monomer without adding precursor substrates. In addition, the 70 L stirrer bioreactor improved P(3HB-<i>co</i>-2 mol% 3HHx) yield 1.4-fold over the flask scale without altering monomer composition. Furthermore, the characterization of copolymer properties showed that this copolymer is promising for skin tissue engineering applications.