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changes create the space in the pocket required to accommodate inhibitors. In other words, significant
conformational changes occur during the process of inhibitor binding that lead to the formation of the
NNIBP [33–35]. This observation also underscores the importance of determining structures of HIV-1
RT with and without bound inhibitors.
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An obvious question is, what forces initiate this series of conformational changes during NNRTI
binding? One possibility is the contacts between the inhibitor and the protein. Though the NNIBP is
hydrophobic, there are three hydrophilic amino acid residues (Lys101 and Lys103 of p66, and Glu138 of
p51) at the rim of the putative entrance(s) to the pocket. The flexible and polar side chains of these
residues could assist in steering an inhibitor into the pocket and/or could block the bound inhibitor from
escaping out of the pocket. Mutagenesis studies have shown that these three residues are important in
the binding of NNRTIs. Though the importance could be explained in terms of the interactions between
these residues and the bound inhibitor in the final complexes, interactions at the initial stages of inhibitor
binding might also be crucial. The flexible and polar side chains of these residues might help in directing
the inhibitor toward the entrance to the pocket via electrostatic interactions, in part by replacing the
original hydrogen bonds between the drug and the solvent molecules. Any initial energy gains from such
polar interactions could potentially be replaced by hydrogen bonds or other types of interactions
between the inhibitor and alternative residues as the inhibitor moves deeper into the binding pocket. In
addition, significant portions of the aromatic rings of both Tyr181 and Tyr188 are exposed at the bottom
of the surface depression and offer the potential for early π-π interactions with the inhibitor. This type of
π-π interaction might also play an important role in the initial approach of inhibitors to the binding
pocket. This hypothesis may provide a kinetic explanation for the ineffectiveness of NNRTIs against
viral strains of HIV-1 that carry nonaromatic amino acids at positions 181 and 188. As the solvated
inhibitor approaches the enzyme and proceeds to enter the binding pocket, most of the water molecules
of solvation are lost. The few water molecules that remain in the NNRTI-bound complex are typically
located at the entrance to the pocket, forming water bridges between the inhibitor and one or two polar
residues around the entrance [33,35,36]. Once the inhibitor is in place, the surface residues close down
around the drug preventing it from escaping by effectively sealing the entrance to the pocket.
VIII. Mechanisms of Inhibition by NNRTIS
Based on structural, biochemical, and genetic data several hypotheses have been postulated about the
mechanism(s) of inhibition of HIV-1 RT by NNRTIs. It is
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