ThePseudomonas aeruginosa liuE gene encodes the 3-hydroxy-3-methylglutaryl coenzyme A lyase, involved in leucine and acyclic terpene catabolism

Abstract

The enzymes involved in the catabolism of leucine are encoded by the liu gene cluster in Pseudomonas aeruginosa PAO1. A mutant in the liuE gene (ORF PA2011) of P. aeruginosa was unable to utilize both leucine/isovalerate and acyclic terpenes as the carbon source. The liuE mutant grown in culture medium with citronellol accumulated metabolites of the acyclic terpene pathway, suggesting an involvement of liuE in both leucine/isovalerate and acyclic terpene catabolic pathways. The LiuE protein was expressed as a His-tagged recombinant polypeptide purified by affinity chromatography in Escherichia coli. LiuE showed a mass of 33 kDa under denaturing and 79 kDa under nondenaturing conditions. Protein sequence alignment and fingerprint sequencing suggested that liuE encodes 3-hydroxy-3-methylglutaryl-coenzyme A lyase (HMG-CoA lyase), which catalyzes the cleavage of HMG-CoA to acetyl-CoA and acetoacetate. LiuE showed HMG-CoA lyase optimal activity at a pH of 7.0 and 37 degrees C, an apparent K(m) of 100 microM for HMG-CoA and a V(max) of 21 micromol min(-1) mg(-1). These results demonstrate that the liuE gene of P. aeruginosa encodes for the HMG-CoA lyase, an essential enzyme for growth in both leucine and acyclic terpenes.