The X-ray crystal structures of 12 substrate-based peptidic inhibitors bound in the active site of the aspartyl protease, HIV-1 protease, have been compared. The inhibitor-binding modes of these inhibitors are remarkably similar despite their structural diversity and conformational flexibility, This prompted the design of a bicyclic peptidomimetic inhibitor 13 with macrocyclic components in constrained conformations that are preorganized for receptor-binding. This inhibitor is a consensus conformational mimic of the protease-bound inhibitor structures with superior properties to peptides, including stability to acid and peptidases as well as antiviral activity. Each of its 15- and 16-membered rings, formed through side-chain to backbone condensation, contains two proteolytically resistant amide bonds and either isoleucine or valine linked via a short aliphatic spacer to tyrosine, The two cycles are connected by a hydroxyethylamine transition state isostere. Molecular modeling and NMR studies indicate that each macrocycle is a highly constrained structural mimic of tripeptide components of linear peptide substrates/inhibitors of HIV-1 protease. Thus the bicyclic peptidomimetic superimposes upon and structurally mimics acyclic hexapeptide inhibitors and their analogues. This results in functional mimicry, as demonstrated by comparable inhibition of HIV-1 protease by acyclic and cyclic molecules at nanomolar concentrations. The rational design of cycles which fix receptor-bound conformations of these bioactive peptides has potential applications for the structural mimicry of other bioactive peptides and may facilitate rational drug design.