Three factors that modulate the activity of class D β-lactamases and interfere with the post-translational carboxylation of Lys70.
The activity of class D β-lactamases is dependent on Lys70 carboxylation in the active site. Structural, kinetic and affinity studies show that this post-translational modification can be affected by the presence of a poor substrate such as moxalactam but also by the V117T substitution. Val117 is a strictly conserved hydrophobic residue located in the active site. In addition, inhibition of class D β-lactamases by chloride ions is due to a competition between the side chain carboxylate of the modified Lys70 and chloride ions. Determination of the individual kinetic constants shows that the deacylation of the acyl-enzyme is the rate-limiting step for the wild-type OXA-10 β-lactamase.
Laboratory of Biological Macromolecules, Centre for Protein Engineering, Institut de Chimie B6a, University of Liège, Liège 4000, Belgium.
Lionel
L
Vercheval
Cédric
C
Bauvois
Alexandre
A
di Paolo
Franck
F
Borel
Jean-Luc
JL
Ferrer
Eric
E
Sauvage
André
A
Matagne
Jean-Marie
JM
Frère
Paulette
P
Charlier
Moreno
M
Galleni
Frédéric
F
Kerff
D000215:Acylation; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020134:Catalytic Domain; D002712:Chlorides / Q000737:chemistry; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004791:Enzyme Inhibitors / Q000737:chemistry; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D008239:Lysine / Q000378:metabolism*; D009070:Moxalactam / Q000378:metabolism; D050505:Mutant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D009994:Osmolar Concentration; D011485:Protein Binding; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D011550:Pseudomonas aeruginosa / Q000201:enzymology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D001618:beta-Lactamases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D002712:Chlorides; D004791:Enzyme Inhibitors; D050505:Mutant Proteins; D011994:Recombinant Proteins; C425107:beta-lactamase OXA-10; D001618:beta-Lactamases; D008239:Lysine; D009070:Moxalactam