Abstract
Staphylococcus aureus is one of the most frequent causes of nosocomial and community-acquired infections, with emerging multiresistant isolates causing a significant burden to public health systems. We identified 2-sulfonylpyrimidines as a new class of potent inhibitors against S. aureus sortase A acting by covalent modification of the active site cysteine 184. Series of derivatives were synthesized to derive structure-activity relationship (SAR) with the most potent compounds displaying low micromolar KI values. Studies on the inhibition selectivity of homologous cysteine proteases showed that 2-sulfonylpyrimidines reacted efficiently with protonated cysteine residues as found in sortase A, though surprisingly, no reaction occurred with the more nucleophilic cysteine residue from imidazolinium-thiolate dyads of cathepsin-like proteases. By means of enzymatic and chemical kinetics as well as quantum chemical calculations, it could be rationalized that the S N Ar reaction between protonated cysteine residues and 2-sulfonylpyrimidines proceeds in a concerted fashion, and the mechanism involves a ternary transition state with a conjugated base. Molecular docking and enzyme inhibition at variable pH values allowed us to hypothesize that in sortase A this base is represented by the catalytic histidine 120, which could be substantiated by QM model calculation with 4-methylimidazole as histidine analog.
Highlights
The emergence of bacterial strains resistant to antibiotic therapy is one of the greatest medical challenges of our time
To evaluate the inhibition potency of the parent sulfonylpyrimidine 4, this compound was tested by means of a fluorometric enzyme assay with recombinantly expressed S. aureus sortase A (SrtA) (Schmohl et al, 2017b) and Abz-LPETG-Dap (Dnp)-OH as substrate
Parent compound 4 was found to act as a time-dependent and irreversible inhibitor, which is in agreement with the literature data (Lit.: 97% inhibition after 16 h of incubation with 100 μM inhibitor; Jaudzems et al, 2020)
Summary
The emergence of bacterial strains resistant to antibiotic therapy is one of the greatest medical challenges of our time. The cysteine transpeptidase sortase A (SrtA) has been discussed as an antivirulence drug target for nearly 20 years since SrtA mediates the attachment of virulence-associated surface proteins to the bacterial cell wall (Perry et al, 2002). Neither genetic deletion (Mazmanian et al, 1999) nor selective chemical inhibition (Cascioferro et al, 2014; Zhang et al, 2014; Mu et al, 2018) of S. aureus SrtA was found to affect the growth properties of bacterial cells, deducing a lower selective pressure for resistance development compared to bactericidal antibiotics. The fact that SrtA plays a key role in the pathogenesis of S. aureus and the enzyme is drug-accessible on the outside of the bacterial cell membrane makes SrtA seemingly a well-druggable target for the development of anti-virulence agents (Cascioferro et al, 2015)
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