Expression, purification, crystallization and preliminary X-ray analysis of the native class C beta-lactamase from Enterobacter cloacae 908R and two mutants.
Crystals have been obtained of the Enterobacter cloacae 908R beta-lactamase and two point mutants by the vapour-diffusion method using similar conditions [pH 9.0, polyethylene glycol (M(r) = 6000) as precipitant]. The three crystal forms belong to the orthorhombic space group P2(1)2(1)2, with roughly the same unit-cell parameters; i.e. for the wild-type crystals a = 46.46, b = 82.96, c = 95.31 A. In the best cases, the crystals diffract to about 2.1 A resolution on a rotating-anode X-ray source at room temperature. Co-crystallization experiments of poor substrates with the wild-type protein and the active-site serine mutant (S64C) are planned and should lead to a better understanding of the catalytic mechanism of class C beta-lactamases.
Wouters
J
J
Laboratoire de Chimie Moléculaire Structurale, Facultés Universitaires Notre Dame de la Paix, 61 Rue de Bruxelles, B-5000 Namur, Belgium. johan.wouters@fundp.ac.be
Charlier
P
P
Monnaie
D
D
Frère
J M
JM
Fonzé
E
E
D003460:Crystallization; D018360:Crystallography, X-Ray; D016972:Enterobacter cloacae / Q000201:enzymology* / Q000235:genetics; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D001618:beta-Lactamases / Q000737:chemistry* / Q000235:genetics
D011994:Recombinant Proteins; D001618:beta-Lactamases