Expression, purification, crystallization and preliminary X-ray analysis of the native class C beta-lactamase from Enterobacter cloacae 908R and two mutants.
[en] 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.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Wouters, J.
Charlier, Paulette ; Université de Liège - ULiège > Département des sciences de la vie > Cristallographie des macromolécules biologiques
Monnaie, D.
Frère, Jean-Marie ; Université de Liège - ULiège > Centre d'ingénierie des protéines
Fonze, E.
Language :
English
Title :
Expression, purification, crystallization and preliminary X-ray analysis of the native class C beta-lactamase from Enterobacter cloacae 908R and two mutants.
Bush K., Jacoby G.A., Medeiros A.A. Antimicrob. Agents Chemother. 1995, 39:1211-1233.
Cantor C.R., Schimmel P.R. Biophys. Chem. 1980, 2:380-381.
Acta Cryst. 1994, 50 D:760-763.
Crichlow G.V., Kuzin A.P., Nugaka M., Mayama K., Sawai T., Knox J.R. Biochemistry 1999, 38:10256-10261.
Dubus A., Monnaie D., Jacobs C., Normark S., Frère J.-M. Biochem. J. 1993, 292:537-543.
Frere J.M. Mol. Microbiol. 1995, 16:385-395.
Galleni M., Amicosante G., Frère J.-M. Biochem. J. 1988, 255:123-129.
Grasseti D.R., Murray J.F. Jr. Arch. Biochem. Biophys. 1967, 119:41-49.
Heinze-Krauss I., Angehrn P., Charnas R.L., Gubernator K., Gutknecht E.M., Hubschwerlen C., Kania M., Oefner C., Page M.G., Sogabe S., Specklin J.L., Winkler F. J. Med. Chem. 1998, 41:3961-3971.
Howard A.J., Gilliland G.L., Finzel B.C., Poulos T., Ohlendorf D.O., Salemme F.R. J. Appl. Cryst. 1987, 20:383-387.
Jacobs C., Dubus A., Monnaie D., Normark S., Frère J.-M. FEMS Microbiol. Lett. 1992, 92:95-100.
Lindberg F., Normark S. J. Bacteriol. 1987, 169:758-763.
Sambrook J., Fritsch E.F., Maniatis T. Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press; 1989.