Predictive analysis of ceftazidime hydrolysis in CTX-M-type β-lactamase family members with a mutational substitution at position 167

The CTX-M family of extended-spectrum β-lactamases has been increasing in number over recent years. Its members preferentially hydrolyse cefotaxime over ceftazidime. Recently, ceftazidime-hydrolysing CTX-M β-lactamase producers with a mutation at Pro167Ser have been found. The aim of this study was to determine whether members of the CTX-M-type β-lactamase family are capable of ceftazidime hydrolysis after introduction of the Pro167Ser point mutation. MICs of wild-type enzyme producers for cefotaxime were 2–4 times higher than those of their respective Pro167Ser mutants, whereas MICs of wild-type enzyme producers for ceftazidime were 4–32 times lower than those of their respective Pro167Ser mutants. The kcat/Km values for Pro167Ser mutants and their respective wild-type enzymes were identical for cefalothin, penicillin and nitrocefin. For cefotaxime, catalytic efficiency (kcat/Km) for wild-type enzymes was 3.13–7.12 times higher than that of their respective Pro167Ser mutants. As these enzymes exhibit a very high Km value (>680 mM) for ceftazidime, we measured initial hydrolysis rates for each enzyme at a low substrate concentration (10 μM) to obtain their kcat and kcat/Km values. Under these conditions, Pro167Ser mutants had kcat/Km values 1.73–2.21 times higher than those of their respective wild-type enzymes. These results indicate that the CTX-M-type β-lactamase family can hydrolyse ceftazidime more efficiently because of the point mutation at position 167.