Identification, crystallization and preliminary X-ray diffraction analysis of esterase A from Caulobacter crescentus CB15, a family VIII lipolytic enzyme.

ABSTRACT: The structures and functions of family VIII lipolytic enzymes, which have moderate sequence identity to class C ?-lactamases and penicillin-binding proteins, are largely unknown. Here, the X-ray crystallographic study of a family VIII esterase from Caulobacter crescentus CB15 (CcEstA) is described. Sequence analysis revealed that CcEstA has a conserved serine residue within the S-X-X-K motif which acts as a catalytic nucleophile. Recombinant protein containing an N-terminal His tag was expressed in Escherichia coli and purified to homogeneity. Functional studies showed that CcEstA acts on ?- and ?-naphthyl acetate as substrates. In addition, it can catalyze the hydrolysis of ketoprofen ethyl ester, a highly useful product in industrial applications. CcEstA was crystallized using a solution consisting of 1.0?M potassium/sodium tartrate, 0.1?M imidazole pH 8.0, 0.2?M NaCl, and X-ray diffraction data were collected to a resolution of 1.62?Å with an R(merge) of 9.4%. The crystals of CcEstA belonged to space group C222(1), with unit-cell parameters a = 172.23, b = 176.68, c = 47.93?Å. Structure determination is in progress.

SUBMITTER: Ryu BH 

PROVIDER: S-EPMC3374513 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature