Oxazolidinones mechanism of action: Inhibition of the first peptide bond formation

Oxazolidinones are potent inhibitors of bacterial protein biosynthesis. Pre vious studies have demonstrated that this new class of antimicrobial agent blocks translation by inhibiting initiation complex formation, while post-i nitiation translation by polysomes and poly(U)-dependent translation is not a target for these compounds. We found that oxazolidinones inhibit transla tion of natural mRNA templates but have no significant effect on poly(A)-de pendent translation. Here we show that various oxazolidinones inhibit ribos omal peptidyltransferase activity in the simple reaction of 70 S ribosomes using initiator-tRNA or N-protected CCA-Phe as a P-site substrate and purom ycin as an A-site substrate. Steady-state kinetic analysis shows that oxazo lidinones display a competitive inhibition pattern with respect to both the P-site and A-site substrates. This is consistent with a rapid equilibrium, ordered mechanism of the peptidyltransferase reaction, wherein binding of the A-site substrate can occur only after complex formation between peptidy ltransferase and the P-site substrate. We propose that oxazolidinones inhib it bacterial protein biosynthesis by interfering with the binding of initia tor fMet-tRNA(i)(Met) to the ribosomal peptidyltransferase P-site, which is vacant only prior to the formation of the first peptide bond.