Class I and III polyhydroxyalkanoate synthases from Ralstonia eutropha andAllochromatium vinosum: Characterization and substrate specificity studies

Class I and III polyhydroxyalkanoate (PHA) synthases catalyze the conversio n of beta -hydroxybutyryl coenzyme A (HBCoA) to polyhydroxybutyrate. The Cl ass I PHA synthase from Ralstonia eutropha has been purified by numerous la bs with reported specific activities that vary between 1 and 160 U/mg. An N -terminal (His)(6)-PHA synthase was constructed and purified with specific activity of 40 U/mg. The variable activity is shown to be related to the pr otein's propensity to aggregate and not to incomplete post-translational mo dification by coenzyme A and a phosphopantetheinyl transferase. The substra te specificities of this enzyme and the Class III PRA synthase from Allochr omatium vinosum have been determined with nine analogs of varied chain leng th and branching, OH group position within the chain, and thioesters. The r esults suggest that in vitro, both PRA synthases are very specific and prov ide further support for their active site structural similarities. In vitro results differ from studies in vivo. (C) 2001 Academic Press.