THE STRUCTURE-BASED DESIGN OF THREE POTENT INHIBITORS OF TEM-1 [[beta]]-LACTAMASE. Natalie C. J. Strynadka^l, Richard Martin³, Bryan Jones³, John Vederas², Michael N. G James^l, ¹MRC Group in Protein Structure and Function, Department of Biochemistry, ²Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada, ³Department of Chemistry, University of Toronto, Toronto, Canada

TEM-1 [[beta]]-lactamase is a wide-spread plasmid-mediated enzyme responsible for inducing antibiotic resistance in bacterial species via its ability to cleave and inactivate the beta-lactam ring in the classic penicillin and cephalosporin families of antibiotic drugs. TEM-1 is particularily notorious in the clinical setting in that newly arising mutants have shown resistance to an ever-increasing spectrum of [[beta]]-lactam drugs as well as the small number of currently existing [[beta]]-lactam inhibitors[l].

We have used the high-resolution structural coordinates of native TEM-1 [2], of the acyl-enzyme intermediate complex of TEM- 1 and the substrate Penicillin G [2], and of the complex of TEM-1 with a large, 165 amino-acid [[beta]]-lactamase inhibitory protein, BLIP [3,4], to design three novel, small-molecule inhibitors against TEM-1. The first two compounds were designed to mimic transition state intermediates in the reaction pathway. These molecules have been synthesized and analyzed kinetically for their ability to inhibit TEM-1 [[beta]]-lactamase. They are shown to be highly potent both against the enzyme and bacterial cells in culture with K_i's in the nM range. The third inhibitor is a non-peptide analog of the beta-hairpin of BLIP that was observed to inhibit the active site of TEM-1 in the TEM-1/BLIP complex[4} .

The structures of each of the enzyme-inhibitor complexes has been solved and refined to 1.7 Å resoluiton. The details of the design strategy and the resulting kinetic and strucutral observations will be discussed.

1. Blazquez, J. et al. Antimicrob. Agents. Chemother. 37, 2059-67 (1993)

2. Strynadka, N. C. J., et al. Nature 359, 700-705 (1992)

3. Strynadka, N. C. J., et al., Nature 368, 657-660. (1994)

4. Strynadka, N. C. J., et al., Nature:Structural Biology (in press)