This proposal responds to the growing problem of antibiotic resistance in the institutionalized elderly. Infection by Gram negative organisms is a major cause of morbidity and mortality in this population. In the nursing home, these infections are often treated empirically using costly extended spectrum (third generation) cephalosporins or beta-lactam beta- lactamase inhibitor combinations (amoxacillin/clavulanic acid). An inevitable result of this strategy is the continued selection of microorganisms with plasmid mediated beta-lactamases that are resistant to these agents. This evolution will ultimately compromise our ability to manage patients with infections in the nursing home. In many instances, the molecular basis of this resistance is a single amino acid change in the beta-lactamase enzyme. Such changes either improve the enzyme's ability to inactivate the antibiotic before it reaches its target or render it resistant to inactivation by the beta-lactamase inhibitors. The central aim of this project is to understand why certain amino acid substitutions dramatically alter beta-lactamase activity and how these changes compare with other common beta-lactamases. We have identified the OHIO-1 beta-lactamase as an excellent model for study, since the entire spectrum of known beta-lactamase variants have been identified in the class A enzyme. We propose to purify and kinetically characterize the native and mutant enzymes of OHIO-1 beta-3-lactamase, and carry out molecular modeling. A detailed understanding of these and related beta-lactamases will permit the rational design of treatment strategies that will minimize the emergence of resistance in these settings.
|Hujer, Andrea M; Hujer, Kristine M; Helfand, Marion S et al. (2002) Amino acid substitutions at Ambler position Gly238 in the SHV-1 beta-lactamase: exploring sequence requirements for resistance to penicillins and cephalosporins. Antimicrob Agents Chemother 46:3971-7|
|Hujer, A M; Hujer, K M; Bonomo, R A (2001) Mutagenesis of amino acid residues in the SHV-1 beta-lactamase: the premier role of Gly238Ser in penicillin and cephalosporin resistance. Biochim Biophys Acta 1547:37-50|
|Bonomo, R A; Knox, J R; Rudin, S D et al. (1997) Construction and characterization of an OHIO-1 beta-lactamase bearing Met69Ile and Gly238Ser mutations. Antimicrob Agents Chemother 41:1940-3|
|Bonomo, R A; Rudin, S D; Shlaes, D M (1997) OHIO-1 beta-lactamase mutants: Asp179Gly mutation confers resistance to ceftazidime. FEMS Microbiol Lett 152:275-8|