Pneumococcal beta-lactam resistance due to a conformational change in penicillin-binding protein 2x.
Streptococcus pneumoniae is a life-threatening human pathogen that is increasingly resistant to a wide array of drugs. Resistance to beta-lactams, the most widely used antibiotics, is correlated with tens of amino acid substitutions in their targets; that is, the penicillin-binding proteins (PBPs), resulting from multiple events of recombination. To discriminate relevant substitutions from those that are incidental to the recombination process, we report the exhaustive characterization of all the mutations in the transpeptidase domain of PBP2x from the highly resistant strain 5204. A semi-automated method combining biochemical and microbiological approaches singled out 6 mutations of 41 (15%) that are essential for high level resistance. The hitherto uncharacterized I371T, R384G, M400T, and N605T together with the previously studied T338M and M339F account for nearly all the loss of affinity of PBP2x for beta-lactams. Most interestingly, I371T and R384G cause the conformational change of a loop that borders the entrance of the active site cavity, hampering antibiotic binding. For the first time all the mutations of a PBP relevant to beta-lactam resistance have been identified, providing new mechanistic insights. Most notable is the relationship between the decreased susceptibility to beta-lactams and the dynamic behavior of a loop.
Carapito
Raphaël
R
Laboratoire d'Ingénierie des Macromolécules, Institut de Biologie Structurale (Commissariat à l'Energie Atomique/CNRS UMR 5075/Université Joseph Fourrier), Grenoble, France.
Chesnel
Laurent
L
Vernet
Thierry
T
Zapun
André
A
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002439:Cefotaxime / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D046915:Penicillin-Binding Proteins / Q000737:chemistry / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D017433:Protein Structure, Secondary; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013296:Streptococcus pneumoniae / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D018440:beta-Lactam Resistance / Q000502:physiology*
D046915:Penicillin-Binding Proteins; C072223:PBP 2x protein, Streptococcus; D002439:Cefotaxime