Thiomandelic acid, a broad spectrum inhibitor of zinc beta-lactamases: kinetic and spectroscopic studies.
Resistance to beta-lactam antibiotics mediated by metallo-beta-lactamases is an increasingly worrying clinical problem. Candidate inhibitors include mercaptocarboxylic acids, and we report studies of a simple such compound, thiomandelic acid. A series of 35 analogues were synthesized and examined as metallo-beta-lactamase inhibitors. The K(i) values (Bacillus cereus enzyme) are 0.09 microm for R-thiomandelic acid and 1.28 microm for the S-isomer. Structure-activity relationships show that the thiol is essential for activity and the carboxylate increases potency; the affinity is greatest when these groups are close together. Thioesters of thiomandelic acid are substrates for the enzyme, liberating thiomandelic acid, suggesting a starting point for the design of "pro-drugs." Importantly, thiomandelic acid is a broad spectrum inhibitor of metallo-beta-lactamases, with a submicromolar K(i) value for all nine enzymes tested, except the Aeromonas hydrophila enzyme; such a wide spectrum of activity is unprecedented. The binding of thiomandelic acid to the B. cereus enzyme was studied by NMR; the results are consistent with the idea that the inhibitor thiol binds to both zinc ions, while its carboxylate binds to Arg(91). Amide chemical shift perturbations for residues 30-40 (the beta(3)-beta(4) loop) suggest that this small inhibitor induces a movement of this loop of the kind seen for other larger inhibitors.
Mollard
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C
Biological NMR Centre, Department of Biochemistry, University of Leicester, P.O. Box 138, University Rd., Leicester LE1 9HN, United Kingdom.
Moali
C
C
Papamicael
C
C
Damblon
C
C
Vessilier
S
S
Amicosante
G
G
Schofield
C J
CJ
Galleni
M
M
Frere
J M
JM
Roberts
G C
GC
D001120:Arginine / Q000737:chemistry; D001665:Binding Sites; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008333:Mandelic Acids / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D008956:Models, Chemical; D008958:Models, Molecular; D011485:Protein Binding; D013053:Spectrophotometry; D013329:Structure-Activity Relationship; D013438:Sulfhydryl Compounds / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry / Q000378:metabolism*; D065093:beta-Lactamase Inhibitors
D004791:Enzyme Inhibitors; D008333:Mandelic Acids; D013438:Sulfhydryl Compounds; D065093:beta-Lactamase Inhibitors; C443128:thiomandelic acid; D001120:Arginine; D015032:Zinc