[en] The lysine-234 residue is highly conserved in beta-lactamases and in nearly all active-site-serine penicillin-recognizing enzymes. Its replacement by a histidine residue in the Streptomyces albus G class A beta-lactamase yielded an enzyme the pH-dependence of which was characterized by the appearance of a novel pK, which could be attributed to the newly introduced residue. At low pH, the k(cat.) value for benzylpenicillin was as high as 50 % of that of the wild-type enzyme, demonstrating that an efficient active site was maintained. Both k(cat.) and k(cat.)/K(m) dramatically decreased above pH 6 but the decrease in k(cat.)/K(m) could not be attributed to larger K(m) values. Thus a positive charge on the side chain of residue 234 appears to be more essential for transition-state stabilization than for initial recognition of the substrate ground state.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
BRANNIGAN, J.; University of Sussex
Matagne, André ; Université de Liège - ULiège > Laboratoire d'Enzymologie, Centre d'Ingénierie des Protéines
Jacob, Françoise
Damblon, Christian ; Université de Liège - ULiège > Laboratoire d'Enzymologie, Centre d'Ingénierie des Protéines
Joris, Bernard ; Université de Liège - ULiège > Laboratoire d'Enzymologie, Centre d'Ingénierie des Protéines
KLEIN, D.; Université de Liège - ULiège > Laboratoire d'Enzymologie, Centre d'Ingénierie des Protéines
SPRATT, B. G.; University of Sussex
Frère, Jean-Marie ; Université de Liège - ULiège > Laboratoire d'Enzymologie, Centre d'Ingénierie des Protéines
Language :
English
Title :
THE MUTATION LYS234HIS YIELDS A CLASS-A BETA-LACTAMASE WITH A NOVEL PH-DEPENDENCE