The HIV-1 protease (PR) mediates its own release (autoprocessing) from the polyprotein precursor, Gag-Pol, flanked by the transframe region (TFR) and reverse transcriptase at its N- and C-termini, respectively. Autoprocessing at the N-terminus of PR mediates stable dimer formation essential for catalytic activity, leading to the formation of infectious virus. An antiparallel β-sheet interface formed by the four N- and C-terminal residues of each subunit is important for dimer stability. Here, we present the first high-resolution crystal structures of model protease precursor-clinical inhibitor (PI darunavir or saquinavir) complexes, revealing varying conformations of the N-terminal flanking (S(-4)FNF(-1)) and interface residues (P(1)QIT(4)). A 180° rotation of the T(4)-L(5) peptide bond is accompanied by a new Q(2)-L(5) hydrogen bond and complete disengagement of PQIT from the β-sheet dimer interface, which may be a feature for intramolecular autoprocessing. This result is consistent with drastically lower thermal stability by 14-20 °C of PI complexes of precursors and the mature PR lacking its PQIT residues (by 18.3 °C). Similar to the TFR-PR precursor, this deletion also results in a darunavir dissociation constant (2 × 10(4))-fold higher and a markedly increased dimer dissociation constant relative to the mature PR. The terminal β-sheet perturbations of the dimeric structure likely account for the drastically poorer inhibition of autoprocessing of TFR-PR relative to the mature PR, even though significant differences in active site-PI interactions in these structures were not observed. The novel conformations of the dimer interface may be exploited to target selectively the protease precursor prior to its N-terminal cleavage.