Interaction energies between beta-lactam antibiotics and E-coli penicillin-binding protein 5 by reversible thermal denaturation

Citation
Bm. Beadle et al., Interaction energies between beta-lactam antibiotics and E-coli penicillin-binding protein 5 by reversible thermal denaturation, PROTEIN SCI, 10(6), 2001, pp. 1254-1259
Citations number
23
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
PROTEIN SCIENCE
ISSN journal
0961-8368 → ACNP
Volume
10
Issue
6
Year of publication
2001
Pages
1254 - 1259
Database
ISI
SICI code
0961-8368(200106)10:6<1254:IEBBAA>2.0.ZU;2-A
Abstract
Penicillin-binding proteins (PBPs) catalyze the final stages of bacterial c ell wall biosynthesis. PBPs form stable covalent complexes with beta -lacta m antibiotics, leading to PBP inactivation and ultimately cell death. To un derstand more clearly how PBPs recognize beta -lactam antibiotics, it is im portant to know their energies of interaction. Because beta -lactam antibio tics bind covalently to PBPs, these energies are difficult to measure throu gh binding equilibria. However, the noncovalent interaction energies betwee n beta -lactam antibiotics and a PBP can be determined through reversible d enaturation of enzyme-antibiotic complexes. Escherichia coti PBP 5, a D-ala nine carboxypeptidase, was reversibly denatured by temperature in an appare ntly two-state manner with a temperature of melting (T-m) of 48.5 degreesC and a van't Hoff enthalpy of unfolding (DeltaH(VH)) of 193 kcal/mole. The b inding of the beta -lactam antibiotics cefoxitin, cloxacillin, moxalactam, and imipenem all stabilized the enzyme significantly, with DeltaT(m) values as high as +4.6 degreesC (a noncovalent interaction energy of +2.7 kcal/mo le). interestingly, the noncovalent interaction energies of these ligands d id not correlate with their second-order acylation rate constants (k(2)/K') . These rate constants indicate the potency of a covalent inhibitor, but th ey appear to have Little to do with interactions within covalent complexes, which is the state of the enzyme often used for structure-based inhibitor design.