Identification, crystallization and preliminary X-ray diffraction analysis of esterase A from Caulobacter crescentus CB15, a family VIII lipolytic enzyme.
The structures and functions of family VIII lipolytic enzymes, which have moderate sequence identity to class C β-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 α- and β-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.
Department of Molecular Science and Technology, Graduate School of Interdisciplinary Programs, Ajou University, Suwon 443-749, Republic of Korea.
Bum Han
BH
Ryu
Tri Duc
TD
Ngo
Eunjin
E
Jang
Sungsoo
S
Kim
Hansol
H
Ju
Kyeong Kyu
KK
Kim
T Doohun
TD
Kim
D000595:Amino Acid Sequence; D016935:Caulobacter crescentus / Q000201:enzymology*; D003460:Crystallization; D018360:Crystallography, X-Ray; D004950:Esterases / Q000737:chemistry*; D008969:Molecular Sequence Data; D016415:Sequence Alignment
D004950:Esterases