A single amino acid substitution at the HIV-1 protease termini dimer interface significantly reduces viral particles processing efficiency

The dimeric form of HIV-1 protease (PR) is required for its full proteolytic activity. The stability of the dimer primarily depends on the termini interface, with N-terminal residues 1–4 of one monomer encountering C-terminal residues 96–99 of another. We made an alanine substitution for valine 3 (V3) or leucine 97 (L97) at the termini dimer interface and tested their proteolytic activity. We found that an alanine substitution for L97 (PRL97A) completely inhibited the proteolytic activity of the PR. However, an alanine substitution for V3 (PRV3A) partially impaired the proteolytic activity. We then introduced two forced-dimerization systems involving nucleocapsid (NC) replacement or the addition of 1-2 leucine zippers to determine whether the proteolytic activity of dimer-defective PRs could be restored. We found that two forced-dimerization systems compensated for the defect in PRV3A, but not in PRL97A. This implies that PRV3A and PRL97A potentially impair the PR via different mechanisms or cause defects in PR activity to different extents. These novel findings will likely serve as a foundation for developing new PR inhibitors for treating drug-resistant HIV-1 infections in the future.