[The pH-dependent catalytic reaction of penicillin G acylase and its mutants].
The pH-dependence in the catalytic reaction of recombinant penicillin G acylase and its mutants from B.megaterium has been studied by using kinetic methods. pK(1) and pK(2)of the residues of the wild type penicillin G a cylase, involved in the catalyzed reaction, were 5.50-5.87 and 10.73, respectively, from the curves of logV(m) and log(V(m)/K(m)) versus pH. Results showed tha t the pK(1) and pK(2) values of these residues of the mutants were similar to that of the wild type. pK(1) of 5.64-5.86 for mutant A and 5.69-6.96 for mutant B were obtained, while pK(2) was 10.61 and 10.48 for mutant A and B, respectively. At the same time, pK values at different temperatures were investigated. The ionization enthalpies(deltaH) were 44.38-59.03 kJ/mol and 147.37 kJ/mol, respectively, from th e curve of pK versus temperature. It was presumed according to the results mentioned above that the ionizing residues, involved in the reaction, wer e histidine and lysine that are localized around the active site.
Chen
Jian-Bo
JB
Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biologi c al Sciences, the Chinese Academy of Sciences, Shanghai 200031, China.
Yang
Sheng
S
Wu
Xing-Jia
XJ
Li
Shi-Yun
SY
Yuan
Zhong-Yi
ZY
D001410:Bacillus megaterium / Q000201:enzymology* / Q000235:genetics; D002384:Catalysis; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D009154:Mutation; D010399:Penicillin Amidase / Q000235:genetics / Q000378:metabolism*; D013696:Temperature
D010399:Penicillin Amidase