Vega_1999_Int.Biodeterior.Biodegradation_43_49

A polyester polyurethane (PU)-degrading enzyme, PU esterase, derived from Pseudomonas fluorescens, a bacterium that utilizes polyester PU as the sole carbon source, was purified to homogeneity as indicated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This enzyme was a soluble, extracellular protein with a molecular mass of 48 kDa and was inhibited by phenylmethylsulfonylfluoride (PMSF). A genomic library of Ps.fluorescens was constructed using the Escherichia coli bacteriophage l vector lZAPII. A recombinant phage exhibiting activity against Impranil DLN was isolated. The gene encoding the polyurethanase (PUase) protein was subcloned into a plasmid expression vectorpT7-6 and expressed in E. coli. Upon expression, the PUase was secreted by the host, displayed esterase activity which was inhibited by PMSF, and in vivo 35S-methionine labeling of the gene product encoded by the open reading frame of the clone insert revealed a single polypeptide with a molecular mass of 48 kDa.