Structural features of cephalosporin acylase reveal the basis of autocatalytic activation.
Cephalosporin acylase (CA), a member of the N-terminal nucleophile hydrolase family, is activated through two steps of intramolecular autoproteolysis, the first mediated by a serine residue, and the second by a glutamate, which releases the pro-segment and produces an active enzyme. In this study, we have determined the crystal structures of mutants which could affect primary or secondary auto-cleavage and of sequential intermediates of a slow-processing mutant at 2.0-2.5A resolutions. The pro-segments of the mutants undergo dynamic conformational changes during activation and adopt surprisingly different loop conformations from one another. However, the autoproteolytic site was found to form a catalytically competent conformation with a solvent water molecule, which was essentially conserved in the CA mutants.
Cho
Ki Joon
KJ
Department of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Republic of Korea.
Kim
Jin Kwang
JK
Lee
Ji-Hye
JH
Shin
Hye Jeong
HJ
Park
Sung Soo
SS
Kim
Kyung Hyun
KH
D002384:Catalysis; D018360:Crystallography, X-Ray; D004789:Enzyme Activation; D009154:Mutation; D010399:Penicillin Amidase / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation; D011549:Pseudomonas / Q000201:enzymology*
D010399:Penicillin Amidase