Molecular Basis of Influence of A501X Mutations in Penicillin-Binding Protein 2 of Neisseria gonorrhoeae Strain 35/02 on Ceftriaxone Resistance.

The increase in the resistance of mutant strains of Neisseria gonorrhoeae to the antibiotic ceftriaxone is pronounced in the decrease in the second-order acylation rate constant, k₂/K_S, by penicillin-binding protein 2 (PBP2). These changes can be caused by both the decrease in the acylation rate constant, k₂, and the weakening of the binding affinity, i.e., an increase in the substrate constant, K_S. A501X mutations in PBP2 affect second-order acylation rate constants. The PBP2^A501V variant exhibits a higher k₂/K_S value, whereas for PBP2^A501R and PBP2^A501P variants, these values are lower. We performed molecular dynamic simulations with both classical and QM/MM potentials to model both acylation energy profiles and conformational dynamics of four PBP2 variants to explain the origin of k₂/K_S changes. The acylation reaction occurs in two elementary steps, specifically, a nucleophilic attack by the oxygen atom of the Ser310 residue and C–N bond cleavage in the β-lactam ring accompanied by the elimination of the leaving group of ceftriaxone. The energy barrier of the first step increases for PBP2 variants with a decrease in the observed k₂/K_S value. Submicrosecond classic molecular dynamic trajectories with subsequent cluster analysis reveal that the conformation of the β₃–β₄ loop switches from open to closed and its flexibility decreases for PBP2 variants with a lower k₂/K_S value. Thus, the experimentally observed decrease in the k₂/K_S in A501X variants of PBP2 occurs due to both the decrease in the acylation rate constant, k₂, and the increase in K_S. [ABSTRACT FROM AUTHOR]