Controlled production of a polyhydroxyalkanoate (PHA) tetramer containing different mole fraction of 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3 HV), 4 HV and 5 HV units by engineered Cupriavidus necator

Polyhydroxyalkanoates (PHAs) are promising alternatives to existing petrochemical-based plastics because of their bio-degradable properties. However, the limited structural diversity of PHAs has hindered their application. In this study, high mole-fractions of Poly (39 mol% 3HB-co-17 mol% 3 HV-co-44 mol% 4 HV) and Poly (25 mol% 3HB-co-75 mol% 5 HV) were produced from 4- hydroxyvaleric acid and 5-hydroxyvaleric acid, using Cupriavidus necator PHB⁻⁴ harboring the gene phaC_BP-M-CPF4 with modified sequences. In addition, the complex toxicity of precursor mixtures was tested, and it was confirmed that the engineered C. necator was capable of synthesizing Poly (32 mol% 3HB-co-11 mol% 3 HV-co-25 mol% 4 HV-co-32 mol% 5 HV) at low mixture concentrations. Correlation analyses of the precursor ratio and the monomeric mole fractions indicated that each mole fractions could be precisely controlled using the precursor proportion. Physical property analysis confirmed that Poly (3HB-co-3 HV-co-4 HV) is a rubber-like amorphous polymer and Poly (3HB-co-5 HV) has a high tensile strength and elongation at break. Poly (3HB-co-3 HV-co-4 HV-co-5 HV) had a much lower glass transition temperature than the co-, terpolymers containing 3 HV, 4 HV and 5 HV. This study expands the range of possible physical properties of PHAs and contributes to the realization of custom PHA production by suggesting a method for producing PHAs with various physical properties through mole-fraction control of 3 HV, 4 HV and 5 HV.