[en] Thiomandelic acid is a simple, broad spectrum, and reasonably potent inhibitor of metallo-beta-lactamases, enzymes that mediate resistance to beta-lactam antibiotics. We report studies by NMR and perturbed angular correlation (PAC) spectroscopy of the mode of binding of the R and S enantiomers of thiomandelic acid, focusing on their interaction with the two metal ions in cadmium-substituted Bacillus cereus metallo-beta-lactamase. The 113Cd resonances are specifically assigned to the metals in the two individual sites on the protein by using 113Cd-edited 1H NMR spectra. Each enantiomer of thiomandelate produces large downfield shifts of both 113Cd resonances and changes in the PAC spectra, which indicate that they bind such that the thiol of the inhibitor bridges between the two metals. For R-thiomandelate, this is unambiguously confirmed by the observation of scalar coupling between Halpha of the inhibitor and both cadmium ions. The NMR and PAC spectra reveal that the two chiral forms of the inhibitor differ in the details of their coordination geometry. The complex with R-thiomandelate, but not that with the S-enantiomer, shows evidence in the PAC spectra of a dynamic process in the nanosecond time regime, the possible nature of which is discussed. The thiomandelate complex of the mononuclear enzyme can be detected only at low metal to enzyme stoichiometry; the relative populations of mononuclear and binuclear enzyme as a function of cadmium concentration provide clear evidence for positive cooperativity in metal ion binding in the presence of the inhibitor, in contrast to the negative cooperativity observed in the free enzyme.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Damblon, Christian ; Université de Liège - ULiège > Département de chimie (sciences) > Chimie biologique structurale
Jensen, Mikael
Ababou, Abdessamad
Barsukov, Igor
Papamicael, Cyril
Schofield, Christopher J
Olsen, Lars
Bauer, Rogert
Roberts, Gordon C K
Language :
English
Title :
The inhibitor thiomandelic acid binds to both metal ions in metallo-beta-lactamase and induces positive cooperativity in metal binding.
Publication date :
2003
Journal title :
Journal of Biological Chemistry
ISSN :
0021-9258
eISSN :
1083-351X
Publisher :
American Society for Biochemistry and Molecular Biology, Baltimore, United States - Maryland
Wang, Z., Fast, W., Valentine, A. M., and Benkovic, S. J. (1999) Curr. Opin. Chem. Biol. 3, 614-622
Kuwabara, S., and Abraham, E. P. (1967) Biochem. J. 103, 27C-30C
Laraki, N., Franceschini, N., Rossolini, G. M., Santucci, P., Meunier, C., de Pauw, E., Amicosante, G., Frere, J. M., and Galleni, M. (1999) Antimicrob. Agents Chemother. 43, 902-906
Payne, D. J. (1993) J. Med. Microbiol. 39, 93-99
Carfi, A., Pares, S., Duee, E., Galleni, M., Duez, C., Frere, J. M., and Dideberg, O. (1995) EMBO J. 14, 4914-4921
Carfi, A., Duee, E., Paul-Soto, R., Galleni, M., Frere, J. M., and Dideberg, O. (1998) Acta Crystallogr. D Biol. Crystallogr. 54, 45-57
Carfi, A., Duée, E., Galleni, M., Frère, J. M., and Dideberg, O. (1998) Acta Crystallogr. D54, 313-323
Concha, N. O., Rasmussen, B. A., Bush, K., and Herzberg, O. (1996) Structure 4, 823-836
Fabiane, S. M., Sohi, M. K., Wan, T., Payne, D. J., Bateson, J. H., Mitchell, T., and Sutton, B. J. (1998) Biochemistry 37, 12404-12411
Fitzgerald, P. M., Wu, J. K., and Toney, J. H. (1998) Biochemistry 37, 6791-6800
Ullah, J. H., Walsh, T. R., Taylor, I. A., Emery, D. C., Verma, C. S., Gamblin, S. J., and Spencer, J. (1998) J. Mol. Biol. 284, 125-136
Concha, N. O., Rasmussen, B. A., Bush, K., and Herzberg, O. (1997) Protein Sci. 6, 2671-2676
Toney, J. H., Fitzgerald, P. M., Grover-Sharma, N., Olson, S. H., May, W. J., Sundelof, J. G., Vanderwall, D. E., Cleary, K. A., Grant, S. K., Wu, J. K., Kozarich, J. W., Pompliano, D. L., and Hammond, G. G. (1998) Chem. Biol. 5, 185-196
Paul-Soto, R., Zeppezauer, M., Adolph, H. W., Galleni, M., Frere, J.-M., Carfi, A., Dideberg, O., Wouter, J., Hemmingsen, L., and Bauer, R. (1999) Biochemistry 38, 16500-16506
Concha, N. O., Janson, C. A., Rowling, P., Pearson, S., Cheever, C. A., Clarke, B. P., Lewis, C., Galleni, M., Frere, J. M., Payne, D. J., Bateson, J. H., and Abdel-Meguid, S. S. (2000) Biochemistry 39, 4288-4298
Galleni, M., Lamotte-Brasseur, J., Rossolini, G. M., Spencer, J., Dideberg, O., and Frere, J. M. (2001) Antimicrob. Agents Chemother. 45, 660-663
Orellano, E. G., Girardini, J. E., Cricco, J. A., Ceccarelli, E. A., and Vila, A. J. (1998) Biochemistry 37, 10173-10180
Crowder, M. W., Walsh, T. R., Banovic, L., Pettit, M., and Spencer, J. (1998) Antimicrob. Agents Chemother. 42, 921-926
Wang, Z., and Benkovic, S. J. (1998) J. Biol. Chem. 273, 22402
Bounaga, S., Laws, A. P., Galleni, M., and Page, M. I. (1998) Biochem. J. 331, 703-711
Paul-Soto, R., Bauer, R., Frere, J. M., Galleni, M., Meyer-Klaucke, W., Nolting, H., Rossolini, G. M., de Seny, D., Hernandez-Valladares, M., Zeppezauer, M., and Adolph, H. W. (1999) J. Biol. Chem. 274, 13242-13249
Mollard, C., Moali, C., Papamicael, C., Damblon, C., Vessilier, S., Amicosante, G., Schofield, C. J., Galleni, M., Frere, J. M., and Roberts, G. C. K. (2001) J. Biol. Chem. 276, 45015-45023
Hemmingsen, L., Damblon, C., Antony, J., Jensen, M., Adolph, H. W., Wommer, S., Roberts, G. C. K., and Bauer, R. (2001) J. Am. Chem. Soc. 123, 10329-10335
Damblon, C., Prosperi, C., Lian, L. Y., Barsukov, I., Paul-Soto, R., Galleni, M., Frère, J. M., and Roberts, G. C. K. (1999) J. Am. Chem. Soc. 121, 11575-11576
Piotto, M., Sudiek, V., and Sklenar, V. (1992) J. Biomol. NMR 2, 661-665
Kuboniwa, H., Grzesiek, S., Delaglio, F., and Bax, A. (1994) J. Biomol. NMR 4, 871-878
McCoy, M. A., and Mueller, L. (1992) J. Magn. Reson. A 101, 122-130
Hemmingsen, L., Bauer, R., Bjerrum, M. L., Zeppezauer, M., Adolph, H. W., Formicka, G., and Cedergren-Zeppezauer, E. (1996) Eur. J. Biochem. 241, 546-551
Bauer, R., Danielsen, E., Hemmingsen, L., Sorensen, M. V., Ulstrup, J., Friis, E. P., Auld, D. S., and Bjerrum, M. J. (1997) Biochemistry 36, 11514-11524
Bauer, R., Danielsen, E., Hemmingsen, L., Bjerrum, M. J., Hansson, O., and Singh, K. (1997) J. Amer. Chem. Soc. 119, 157-162
Bauer, R. (1985) Q. Rev. Biophys. 18, 1-64
Cornell, W. D., Cieplak, P., Bayly, C. I., Gould, I. R., Merz, Jr., K. M., Ferguson, D. M., Spellmeyer, D. C., Fox, T., Caldwell, J. W., and Kollman, P. A. (1995) J. Amer. Chem. Soc. 117, 5179-5197
Dixon, S. L., and Merz, K. M., Jr. (1997) J. Chem. Phys. 107, 879-893
Gogonea, V., and Merz, K. M., Jr. (1999) J. Phys. Chem. A 103, 5171-5188
Morris, G. M., Goodsell, D. S., Halliday, R. S., Hart, W. E., Belew, R. K., and Olson, A. J. (1998) J. Comp. Chem. 19, 1639-1662
Hanessian, S., Mitessier, N., Therrien, E. (2001) J. Comp.-Aided Mol. Design 16, 673-881
Bauer, R., Jensen, S. J., and Nielsen, B. S. (1988) Hyperfine Interact. 39, 203-234
Baleja, J. D., Thanabal, V., and Wagner, G. (1997) J. Biomol. NMR 10, 397-401
de Seny, D., Heinz, U., Wommer, S., Kiefer, M., Meyer-Klaucke, W., Galleni, M., Frere, J. M., Bauer, R., and Adolph, H. W. (2001) J. Biol. Chem. 276, 45065-45078
Wommer, S., Rival, S., Heinz, U., Galleni, M., Frere, J.-M., Franceschini, N., Amicosante, G., Rasmussen, B., Bauer, R., and Adolph, H.-W. (2002) J. Biol. Chem. 277, 24142-24147