The role of β-oxidation of short-chain alkanoates in polyhydroxyalkanoate copolymer synthesis inAzotobacter vinelandiiUWD

Abstract

Valerate and other short-chain, uneven-length fatty acids promoted the formation of the polyhydroxyalkanoate (PHA) copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) in Azotobacter vinelandii UWD growing in glucose medium. The uptake of valerate was inducible, being repressed by acetate but not by glucose. A likely route that would direct valerate into PHA synthesis involved the β-oxidation pathway. The short-chain fatty acids butyrate, valerate, trans.-2-pentenoate, crotonate, hexanoate, heptanoate, and octanoate induced the coordinate production of the β-oxidation enzymes enoyl-CoA hydratase (EGH) and L-(+)-3-hydroxybutyryl-CoA dehydrogenase (HAD).trans-3-Pentenoate was the best inducer of these activities, which suggested that the isomerase of the β-oxidation complex also was present. However, 3-hydroxyacyl-CoA epimerase activity of the β-oxidation complex was not detected. 3-Ketoacyl-CoA thiolase activity was constitutive in A. vinelandii and appeared to associate only loosely with the 73 000 Da ECH–HAD complex. Thus, 3-ketoacyl-CoA, the end product of HAD activity, could be directed into PHA synthesis through acetoacetyl-CoA reductase generating the 3-hydroxyvalerate subunit of the polymer. When valerate was the sole carbon source, the incorporation of valerate into the polymer was normal, but most of the valerate was directed into metabolism and very little PHA was formed. When glucose also was present, the β-oxidation of short-chain alkanoates inhibited the specific activity of acetoacetyl-CoA reductase and 3-ketothiolase and the PHA yield. A model for PHA synthesis was developed that suggests that the use of fatty acids to promote PHA copolymer formation in A. vinelandii will inevitably result in decreased PHA yield.Key words: β-oxidation, poly(β-hydroxyalkanoate) synthesis, short chain fatty acids, regulation.