The Lys234Arg substitution in the enzyme SHV-72 is a determinant for resistance to clavulanic acid inhibition.
The new beta-lactamase SHV-72 was isolated from clinical Klebsiella pneumoniae INSRA1229, which exhibited the unusual association of resistance to the amoxicillin-clavulanic acid combination (MIC, 64 microg/ml) and susceptibility to cephalosporins, aztreonam, and imipenem. SHV-72 (pI 7.6) harbored the three amino acid substitutions Ile8Phe, Ala146Val, and Lys234Arg. SHV-72 had high catalytic efficiency against penicillins (k(cat)/K(m), 35 to 287 microM(-1) x s(-1)) and no activity against oxyimino beta-lactams. The concentration of clavulanic acid necessary to inhibit the enzyme activity by 50% was 10-fold higher for SHV-72 than for SHV-1. Molecular-dynamics simulation suggested that the Lys234Arg substitution in SHV-72 stabilized an atypical conformation of the Ser130 side chain, which moved the O gamma atom of Ser130 around 3.5 A away from the key O gamma atom of the reactive serine (Ser70). This movement may therefore decrease the susceptibility to clavulanic acid by preventing cross-linking between Ser130 and Ser70.
Mendonça
Nuno
N
Antibiotic Resistance Unit, Department of Infectious Diseases, National Institute of Health Dr. Ricardo Jorge Av. Padre Cruz, 1649-016 Lisbon, Portugal.
Manageiro
Vera
V
Robin
Frédéric
F
Salgado
M José
MJ
Ferreira
Eugénia
E
Caniça
Manuela
M
Bonnet
Richard
R
D019943:Amino Acid Substitution; D000658:Amoxicillin / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001120:Arginine / Q000235:genetics; D001398:Aztreonam / Q000494:pharmacology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002511:Cephalosporins / Q000494:pharmacology; D019818:Clavulanic Acid / Q000494:pharmacology*; D003198:Computer Simulation; D015378:Imipenem / Q000494:pharmacology; D007525:Isoelectric Focusing; D007700:Kinetics; D007711:Klebsiella pneumoniae / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D008239:Lysine / Q000235:genetics; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010406:Penicillins / Q000494:pharmacology; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D002511:Cephalosporins; D010406:Penicillins; D065093:beta-Lactamase Inhibitors; D019818:Clavulanic Acid; D015378:Imipenem; D000658:Amoxicillin; D001120:Arginine; D001618:beta-Lactamases; D001398:Aztreonam; D008239:Lysine