Energetic, structural, and antimicrobial analyses of β-lactam side chain recognition by β-lactamases

Background: Penicillins and cephalosporins are among the most widely used and successful antibiotics. The emergence of resistance to these beta -lactams, most often through bacterial expression of beta -lactamases, threatens public health. To understand how beta -lactamases recognize their substrates, it would be helpful to know their binding energies. Unfortunately, these have been difficult to measure because beta -lactams form covalent adducts with beta -lactamases. This has complicated functional analyses and inhibitor design. Results: To investigate the contribution to interaction energy of the key amide (R1) side chain of beta -lactam antibiotics, eight acylglycineboronic acids that bear the side chains of characteristic penicillins and cephalosporins, as well as four other analogs, were synthesized. These transition-state analogs form reversible adducts with serine beta -lactamases. Therefore, binding energies can be calculated directly from K-i values, The K-i values measured span four orders of magnitude against the Group I beta -lactamase AmpC and three orders of magnitude against the Group II beta -lactamase TEM-1. The acylglycineboronic acids have K-i values as low as 20 nM against AmpC and as low as 390 nM against TEM1I. The inhibitors showed little activity against serine proteases, such as chymotrypsin. R1 side chains characteristic of beta -lactam inhibitors did not have better affinity for AmpC than did side chains characteristic of beta -lactam substrates. Two of the inhibitors reversed the resistance of pathogenic bacteria to beta -lactams in cell culture. Structures of two inhibitors in their complexes with AmpC were determined by X-ray Crystallography to 1.90 Angstrom and 1.75 Angstrom resolution; these structures suggest interactions that are important to the affinity of the inhibitors. Conclusions: Acylglycineboronic acids allow us to begin to dissect interaction energies between beta -lactam side chains and beta -lactamases, Surprisingly, there is little correlation between the affinity contributed by R1 side chains and their occurrence in beta -lactum inhibitors or beta -lactam substrates of serine p-lactamases. Nevertheless, presented in acylglycineboronic acids, these side chains can lead to inhibitors with high affinities and specificities, The structures of their complexes with AmpC give a molecular context to their affinities and may guide the design of antiresistance compounds in this series. (C) 2001 Elsevier Science Ltd, All rights reserved.