Crystallization and preliminary X-ray diffraction analysis of glutaryl 7-aminocephalosporanic acid acylase from Pseudomonas sp. GK16

Glutaryl-7-aminocephalosporanic acid acylase from Pseudomonas sp. GK16 produces glutaryl-7-aminocephalosporanic acid, a key intermediate for the synthesis of cephem antibiotics. Sequence alignment suggests that the enzyme may belong to the N-terminal nucleophile hydrolase superfamily including penicillin G acylase. The enzyme is an (αβ)₂ heterotetramer of two nonidentical subunits. These subunits are derived from a nascent precursor polypeptide that is cleaved proteolytically through a two-step autocatalytic process upon folding. The enzyme has been crystallized using the vapor diffusion method. A bipyramidal crystal form was obtained from a solution containing polyethylene glycol (MW 3350) and calcium chloride. Complete diffraction data sets have been collected up to 2.8 A resolution. The crystal is tetragonal with the space group P4₁2₁2 or P4₃2₁2 and the unit cell parameters are a = b = 73.5 A, c = 380.3. Considerations of the possible values of V(m) account for the presence of a tetramer in the asymmetric unit. (C) 2000 Academic Press.