Metabolic engineering of Pseudomonas putida for the production of various types of short-chain-length polyhydroxyalkanoates from levulinic acid

Poly(3-hydroxybutyrate), a short-chain-length polyhydroxyalkanoate (scl-PHA), is considered as a good alternative to conventional synthetic plastics. However, various biopolymers with diverse characteristics are still in demand. In this study, four different types of scl-PHA were successfully produced by engineering levulinic acid (LA) utilization metabolic pathway and expressing heterologous PHA synthase (PhaEC), acetyl-CoA acetyltransferase (PhaA), and acetyl-CoA reductase (PhaB) in Pseudomonas putida EM42. Poly(3-hydroxybutyrate-co-3hydroxyvalerate) [P(3HB-co-3HV)], poly(3-hydroxyvalerate-co-4-hydroxyvalerate) [P(3HV-co-4HV)] and poly (3-hydroxybutyrate-co-3-hydroxyvalerate-co-4-hydroxyvalerate) [P(3HB-co-3HV-co-4HV)] were produced by the natural LA pathway, poly(4-hydroxyvalerate) by lvaAB-deleted LA pathway, and P(3HV-co-4HV) and P(3HBco-3HV-co-4HV) with relatively high 3HV by fadB-deleted LA pathway. PHA with different monomer fractions could be produced using different PHA synthases. Scl-PHA contents reached approximately 40% of cell dry mass under non-optimized flask culture. This demonstrates that the LA catabolic pathway may be a good alternative route to provide monomers for the production of various types of PHA.