2A49

Crystal structure of clavulanic acid bound to E166A variant of SHV-1 beta-lactamase

Summary for 2A49

• Entry DOI: 10.2210/pdb2a49/pdb
• Related: 1rcj 2A3U
• Descriptor: Beta-lactamase SHV-1, N-(2-HYDROXY-4-OXO-BUTYL)-N-(3-OXO-TRANSPROPENYL)AMINE, CYCLOHEXYL-HEXYL-BETA-D-MALTOSIDE, ... (5 entities in total)
• Functional Keywords: beta-lactamase, beta-lactam hydrolase, penicillinase, detergent binding, inhibitor design, covalent intermediate, hydrolase
• Biological source: Klebsiella pneumoniae
• Total number of polymer chains: 1
• Total formula weight: 30732.32

Authors

Padayatti, P.S.,Helfand, M.S.,Totir, M.A.,Carey, M.P.,Carey, P.R.,Bonomo, R.A.,van den Akker, F. (deposition date: 2005-06-28, release date: 2005-08-02, Last modification date: 2024-10-30)

Primary citation

Padayatti, P.S.,Helfand, M.S.,Totir, M.A.,Carey, M.P.,Carey, P.R.,Bonomo, R.A.,van den Akker, F.
High Resolution Crystal Structures of the trans-Enamine Intermediates Formed by Sulbactam and Clavulanic Acid and E166A SHV-1 {beta}-Lactamase.
J.Biol.Chem., 280:34900-34907, 2005

PubMed Abstract: Antibiotic resistance mediated by constantly evolving beta-lactamases is a serious threat to human health. The mechanism of inhibition of these enzymes by therapeutic beta-lactamase inhibitors is probed using a novel approach involving Raman microscopy and x-ray crystallography. We have presented here the high resolution crystal structures of the beta-lactamase inhibitors sulbactam and clavulanic acid bound to the deacylation-deficient E166A variant of SHV-1 beta-lactamase. Our previous Raman measurements have identified the trans-enamine species for both inhibitors and were used to guide the soaking time and concentration to achieve full occupancy of the active sites. The two inhibitor-bound x-ray structures revealed a linear trans-enamine intermediate covalently attached to the active site Ser-70 residue. This intermediate was thought to play a key role in the transient inhibition of class A beta-lactamases. Both the Raman and x-ray data indicated that the clavulanic acid intermediate is decarboxylated. When compared with our previously determined tazobactam-bound inhibitor structure, our new inhibitor-bound structures revealed an increased disorder in the tail region of the inhibitors as well as in the enamine skeleton. The x-ray crystallographic observations correlated with the broadening of the O-C=C-N (enamine) symmetric stretch Raman band near 1595 cm(-1). Band broadening in the sulbactam and clavulanic acid inter-mediates reflected a heterogeneous conformational population that results from variations of torsional angles in the O-(C=O)-C=C=NH-C skeleton. These observations led us to conclude that the conformational stability of the trans-enamine form is critical for their transient inhibitory efficacy.

PubMed: 16055923
DOI: 10.1074/jbc.M505333200

Experimental method

X-RAY DIFFRACTION (1.43 Å)