The effect of the PI side chain on the binding of optimized carboxylate and activated carbonyl inhibitors of the hepatitis C virus NS3 protease

Peptidyl inhibitors of the hepatitis C virus NS3 protease hold much promise as direct-acting antiviral agents against hepatitis C infection. The optimization of N-terminal cleavage products, found to exhibit activity (product inhibition) against the enzyme, has led to potent tripeptide inhibitors that bear free C-terminal carboxylate groups. An analogous activated carbonyl compound (pentafluoroethyl ketone) bearing a PI norvaline (Nva) was found to possess comparable activity against hepatitis C virus protease. However, an analogue bearing an aminocyclopropylcarboxylic acid (Acca) PI residue exhibited very poor activity. F-19-NMR studies indicate that the propensity of the Acca-derived activated carbonyl to form hemiketals is only slightly reduced compared with that of a PI Nva equivalent. These results, as well as molecular modeling studies, argue against steric hindrance of the nucleophilic attack of Ser-139 accounting for the poor mechanism-based inhibition by the former. We hypothesize that the conformational properties of the respective C-termini in the context of an adaptable active site account for the divergent PI structure activity relationships.