Amplification and sequence analysis of phaC gene of polyhydroxybutyrate producing Vibrio azureus BTKB33 isolated from marine sediments
A recombinant of Pseudomonas putida LS461 (deletion of the phaC1phaZphaC2 genes) was constructed by introducing cosmid JC123 carrying a novel phaC116 gene from a metagenomic clone. The resulting strain, P. putida LS46123, was able to synthesize polyhydroxyalkanoate (PHA) polymers with novel monomer compositions when cultured on glucose or free fatty acids, and accumulated PHAs from 9.24% to 27.09% of cell dry weight. The PHAs synthesized by P. putida LS46123 contained up to 50 mol % short chain length subunits (3-hydroxybutyrate and 3-hydroxyvalerate), with the remaining monomers consisting of various medium chain length subunits. The PhaC116 protein expressed by P. putida LS46123 had an amino acid sequence similarity of 45% with the PhaC1 protein of the parent strain, P. putida LS46. Predicted 3D structures of the PhaC116 proteins from P. putida LS46123 and P. putida LS46 revealed several differences in the numbers and locations of protein secondary structures. The physical and thermal properties of the novel polymers synthesized by P. putida LS46123 cultured with glucose or free fatty acids differed significantly from those produced by P. putida LS46 grown on the same substrates. PHA polymers with different subunit compositions, and hence different physical and thermal properties, can be tailor-made using novel PHA synthase for specific applications.