Paper Report for: Pesaresi_2005_Curr.Microbiol_50_102
Reference
Title: Isolation, characterization, and heterologous expression of a carboxylesterase of Pseudomonas aeruginosa PAO1 Pesaresi A, Devescovi G, Lamba D, Venturi V, Degrassi G Ref: Curr Microbiol, 50:102, 2005 : PubMed
We purified to homogeneity an intracellular esterase from the opportunistic pathogen Pseudomonas aeruginosa PAO1. The enzyme hydrolyzes p-nitrophenyl acetate and other acetylated substrates. The N-terminal amino acid sequence was analyzed and 11 residues, SEPLILDAPNA, were determined. The corresponding gene PA3859 was identified in the P. aeruginosa PAO1 genome as the only gene encoding for a protein with this N-terminus. The encoding gene was cloned in Escherichia coli, and the recombinant protein expressed and purified to homogeneity. According to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis and analytical gel filtration chromatography, the esterase was found to be a monomer of approximately 24 kDa. The experimentally determined isoelectric point was 5.2 and the optimal enzyme activity was at 55 degrees C and at pH 9.0. The esterase preferentially hydrolyzed short-chain fatty acids. It is inhibited by phenylmethylsulfonyl fluoride (PMSF) but not by ethylendiaminotetraacetic acid (EDTA). Native enzyme preparations typically showed a Michaelis constant (K(m)) and V(max) of 0.43 mM and 12,500 U mg(-1), respectively, using p-nitrophenyl acetate as substrate. Homology-based database searches clearly revealed the presence of the consensus GXSXG signature motif that is present in the serine-dependent acylhydrolase protein family.
Pesaresi A, Devescovi G, Lamba D, Venturi V, Degrassi G (2005) Isolation, characterization, and heterologous expression of a carboxylesterase of Pseudomonas aeruginosa PAO1 Curr Microbiol50: 102-9
Pesaresi A, Devescovi G, Lamba D, Venturi V, Degrassi G (2005) Curr Microbiol50: 102-9