Τίτλος:
Interactions of oximino-substituted boronic acids and β-lactams with the cmy-2-derived extended-spectrum cephalosporinases cmy- 30 and cmy-42
Γλώσσες Τεκμηρίου:
Αγγλικά
Περίληψη:
CMY-30 and CMY-42 are extended-spectrum (ES) derivatives of CMY-2. ES characteristics are due to substitutions of Gly (CMY- 30) and Ser (CMY-42) for Val211 in theβ-loop. To characterize the effects of 211 substitutions, we studied the interactions of CMY-2, -30, and -42 with boronic acid transition state inhibitors (BATSIs) resembling ceftazidime and cefotaxime, assessed thermal stability of the enzymes in their free forms and in complexes with BATSIs and oximino lactams, and simulated, using molecular dynamics (MD), the CMY-42 apoenzyme and the CMY-42 complexes with ceftazidime and the ceftazidime-like BATSI. Inhibition constants showed that affinities between CMY-30 and CMY-42 and the R1 groups of BATSIs were lower than those of CMY-2. ES variants also exhibited decreased thermal stability either as apoenzymes or in covalent complexes with oximino compounds.MDsimulations further supported destabilization of the ES variants. Val211Ser increased thermal factors of theloop backbone atoms, as previously observed for CMY-30. The similar effects of the two substitutions seemed to be due to a less-constrained Tyr221 likely inducing concerted movement of elements at the edges of the active site (loop-Q120 loop-R2 loop/H10 helix). This inner-protein movement, along with the wider R1 binding cleft, enabled intense vibrations of the covalently bound ceftazidime and ceftazidime-like BATSIs. Increased flexibility of the ES enzymes may assist the productive adaptation of the active site to the various geometries of the oximino substrates during the reaction (higher frequency of near-attack conformations). Copyright © 2013, American Society for Microbiology. All Rights Reserved.
Συγγραφείς:
Kotsakis, S.D.
Caselli, E.
Tzouvelekis, L.S.
Petinaki, E.
Prati, F.
Miriagou, V.
Περιοδικό:
Antimicrobial Agents and Chemotherapy
Λέξεις-κλειδιά:
boronic acid transition state inhibitor; cephalosporinase; extended spectrum cephalosporinase CMY 30; extended spectrum cephalosporinase CMY 42; hydrolase inhibitor; oximino beta lactam; unclassified drug, article; binding affinity; dynamics; enzyme inhibition; enzyme stability; priority journal; thermostability, Amino Acid Substitution; beta-Lactamases; beta-Lactams; Boronic Acids; Cephalosporin Resistance; Cephalosporinase; Cephalosporins; Citrobacter freundii; Escherichia coli; Escherichia coli Proteins; Molecular Dynamics Simulation; Protein Structure, Tertiary
DOI:
10.1128/AAC.01620-12
