PRODUCTION OF HETEROPOLYMERIC POLYHYDROXYALKANOATE IN ESCHERICHIA-COLI FROM A SINGLE CARBON SOURCE

Poly[beta-hydroxybutyrate-co-beta-hydroxyvalerate] co-polymer, PHBV, i s a polyhydroxyalkanoate (PHA) that has greater utility as a biodegrad able thermoplastic polyester than poly-beta-hydroxybutyrate, PHB. In o rder to produce PHBV, a system of pathways is required to produce both hydroxybutyrate (HE) and hydroxyvalerate (HV) monomers from the sourc es of carbon. A working model for conversion of glucose to PHBV via ac etyl- and propionyl-coenzyme A was constructed by expressing the PHA b iosynthesis genes from Alcaligenes eutrophus in Escherichia coli strai n K-12 under novel growth conditions. When 1 mM valine was added to 1% glucose medium, growth ceased and up to 2.5% of the incorporated mono mers were HV; up to 4% were HV when 1 mM threonine was added as well. Threonine dehydratase (TD) converts threonine to alpha-ketobutyrate; T D is required for HV to be incorporated into PHA unless its transamina ted reaction product, alpha-aminobutyrate, is added to the medium. Int racellular alpha-ketobutyrate accumulates when valine is added to the medium because valine, which cannot be metabolized to HV by E. coil st rain K-12, stimulates TD and inhibits acetolactate synthase, In turn, alpha-ketobutyrate is converted to propionyl-CoA by the E. coil pyruva te dehydrogenase complex. This constitutes a defined system of pathway s for synthesis of a heteropolymeric PHA from a single carbon source, which in the future could be transferred to other organisms including plants.