The synthesis of poly-(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] copolymer by recombinant Escherichia coli was studied in the medium containing glucose and valeric acid as carbon sources. A recombinant E. coli strain (fadR atoC) harboring a stable high-copy number plasmid containing the Alcaligenes eutrophus polyhydroxy alkanoate (PHA) biosynthesis genes was constructed for the production of the copolymer P(3HB-co-3HV). Accumulation of acetic acid not only had a detrimental effect on cell growth but also decreased the flux of acetyl-CoA into the P(3HB-co-3HV) biosynthetic pathway. Reducing specific growth rate by increasing the initial acetic acid concentration resulted in enhanced copolymer synthesis due to less accumulation of acetic acid. Initial acetic acid concentration of 50mM was found to be optimal at 20g/l glucose and 20mM valeric acid concentration. The fraction of 3-hydroxyva-lerate (3HV) increased with decreasing growth temperature. The ratios of 3HV to 3HB in the copolymer could be controlled by altering the concentrations of valeric acid and glucose in the medium. Catabolite repression was in part responsible for the inefficient copolymer synthesis. Various nutritional components were examined for their ability to relieve catabolite repression. An addition of oleic acid resulted in threefold increase of the 3HV fraction in the copolymer. An addition of a small amount of tryptone and peptone considerably promoted P(3HB-co-3HV)synthesis.