A study on the relation between poly(3-hydroxybutyrate) depolymerases or oligomer hydrolases and molecular weight of polyhydroxyalkanoates accumulating in Cupriavidus necator H16

Abstract

Cupriavidus necator H16 has nine genes of poly(3-hydroxybutyrate) (PHB) depolymerases or oligomer hydrolases (intracellular PHB mobilization enzymes). In this study, we evaluated the relation between these genes and the accumulation, consumption, and molecular weight of polyhydroxyalkanoates (PHAs) accumulating in strain H16 and in a recombinant C. necator strain, KNK-005, which harbors an NSDG mutant of the PHA synthase gene (phaCAc) from Aeromonas caviae. PhaZ6 had a significant influence on the molecular weight of PHA when palm kernel oil was used as a carbon source. The 005dZ6 strain (ΔphaZ6 mutant of KNK-005) could produce ultra-high-molecular-weight poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) with weight-average molecular weight (Mw) >3.0×106 (approximately double that of KNK-005). Under PHA consumption conditions, deletion of phaZ1 and phaZ2 had a significant and slight attenuating effect, respectively, on the reduction in PHA content of KNK-005 cells. Regardless of the PHA consumption, its Mw did not decrease. Thus, 005dZ126 (the ΔphaZ1ΔphaZ2ΔphaZ6 triple mutant of KNK-005) is a promising strain capable of producing PHBHHx of ultra-high-molecular-weight and barely degrades PHBHHx enzymatically intracellularly. This is the first report examining the relation between intracellular PHB mobilization enzymes and molecular weight of PHAs accumulating in C. necator H16 and the derivatives.