Cloning and characterization of the Pseudomonas sp 61-3 phaG gene involvedin polyhydroxyalkanoate biosynthesis

Pseudomonas sp. 61-3 produces a blend of poly(3-hydroxybutyrate) [P(3HB)] h omopolymer and poly(3-whydroxybutyrate-co-3-hydroxyalkanoates) [P(3HB-co-3H A)] random copolymer consisting of monomeric units of 4-12 carbon atoms fro m sugars. The phaG(Ps) gene encoding (R)-3-hydroxyacyl-acyl carrier protein coenzyme A transferase was cloned from this strain, and homologous express ion of this gene under the control of the lac or the native promoter was in vestigated. Additional copies of the phaG(Ps), gene in Pseudomonas sp. 61-3 led to an increase in both the polyhydroxyalkanoate (PHA) content in the c ells and the fraction of medium-chain-length 3HA units in PHA. Disruption o f the chromosomal phaG(Ps) gene resulted in an increase in the fraction of the 3HB unit in PHA. The site-directed mutagenesis of the phaG(Ps) gene was carried out to investigate the role of a HX4D motif which has been propose d to be related to PhaG activity.