Kinetic interactions of tazobactam with beta-lactamases from all major structural classes.
Tazobactam was shown to be a potent inhibitor of group 1, 2a, 2b, and 2b' beta-lactamases. Extended kinetic studies with class A and C serine beta-lactamases showed that the PC1, TEM-2, and P99 enzymes all were reversibly inhibited prior to inactivation of the enzymes. The CcrA metallo-beta-lactamase was less well inhibited, with a 50% inhibitory concentration at least 3 orders of magnitude less favorable than those for most serine beta-lactamases. The numbers of hydrolytic turnovers of tazobactam before inactivation were 2 for PC1, 125 for TEM-2, 50 for P99, and 4,000 for the CcrA enzyme. In spectral studies, transient intermediates were formed after reaction of tazobactam with the PC1, TEM-2, and CcrA beta-lactamases, corresponding to enzyme-associated intermediates responsible for hydrolysis of tazobactam. Chromophores absorbing at 270 nm (CcrA) and 288 nm (TEM-2 and PC1) were observed for these reaction intermediates. The P99 cephalosporinase formed a stable complex with a UV maximum at 295 nm. Incubation of tazobactam with all of the enzymes resulted in accumulation of a tazobactam reaction product with a short-wavelength absorbance. This product has characteristics similar to those of the major eucaryotic metabolite of tazobactam. Possible reaction mechanisms are presented to explain the findings. In conclusion, both serine-based and metallo-beta-lactamases were irreversibly inactivated by tazobactam following an initial transient inhibition phase.
Medical Research Division, American Cyanamid, Pearl River, New York 10965.
K
K
Bush
C
C
Macalintal
B A
BA
Rasmussen
V J
VJ
Lee
Y
Y
Yang
D001426:Bacterial Proteins / Q000378:metabolism; D002511:Cephalosporins / Q000494:pharmacology; D019818:Clavulanic Acid; D002969:Clavulanic Acids / Q000494:pharmacology; D006868:Hydrolysis; D007525:Isoelectric Focusing; D007700:Kinetics; D008970:Molecular Weight; D010397:Penicillanic Acid / Q000378:metabolism / Q000494:pharmacology*; D013056:Spectrophotometry, Ultraviolet; D000078142:Tazobactam; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000032:analysis / Q000302:isolation & purification
D001426:Bacterial Proteins; D002511:Cephalosporins; D002969:Clavulanic Acids; D065093:beta-Lactamase Inhibitors; D019818:Clavulanic Acid; D010397:Penicillanic Acid; D001618:beta-Lactamases; C056342:beta-lactamase TEM-2; C063912:carbapenemase; C021720:nitrocefin; D000078142:Tazobactam