NORMAL TARGET ENZYMEÂ FUNCTION
Saquinavir is an inhibitor of HIV-1 and HIV-2 proteases which binds to their active sites and stops the proteases from cleaving the synthesized HIV proteins in an infected cell, effectively preventing the production of more viruses and aiding in stopping the disease.
HIV-1 PROTEASE STRUCTURE AND IMPORTANT RESIDUES
HIV protease functions normally by cleaving the viral polyprotein precursor into the proteins needed for assembly with the viral RNA at the cell surface, forming fresh virions. This all occurs inside T cells with the CD4+ antigen, the target of HIV. Each HIV protease monomer is only 99 amino acids, so each monomer is technically only a polypeptide itself if we consider 100 or greater to be a protein!
HIV PROTEASE FUNCTION
The majority of amino acids at which HIV protease cleaves are nonpolar.
Pr55Gag polyprotein produces the main structural proteins of HIV, while Pr160Gag-Pol produces the enzymes necessary for the proliferation of HIV, such as the HIV protease itself, the integrase, and the reverse transcriptase. In their mature state, Pr55Gag and Pr160Gag-Pol continue the viral cycle of HIV.(1) Without treatment, these proteins would continue to propel HIV to proliferate and increase viral load in someone infected with HIV. In the maturation of these proteins, HIV proteases are responsible for cleaving immature Pr55Gag and Pr160Gag-Pol at specific cut sites.
The specific cut sites cleaved by HIV protease are mostly between hydrophobic residues.(2) De Olivera et.al successfully identified the most frequent cleavage sites. They found that >97% of these residues were nonpolar. From this 97%, 77.8% were hydrophobic and 21.5% were small amino acids (mostly prolines with some alanines and rarely glycine).
Figure 2. Diagram of where HIV protease cuts, which is primarily at hydrophobic residues.
(Image distributed under the Creative Commons License, see (3))​
MECHANISM OF PROTEIN ACTION
There are two major mechanisms that have been proposed for the peptide bond cleaving action of HIV-1 protease.
HIV protease is an aspartic protease with the conserved sequence D-T-G. If the aspartic acid is mutated to an alanine, then the protease fails to function. Furthermore, the D25 residue of both monomers in this conserved sequence appears to be the catalytic residue.(3) The general aspartic protease mechanism involves the creation of a hydroxide nucleophile by an aspartate accepting a proton from water. This hydroxide attacks the carbonyl carbon of the amide bond, forming an acyl intermediate stabilized by Zn2+. The negative oxygen reforms the carbonyl, and the bond between the carbon and the nitrogen is broken as the nitrogen acts as a base, abstracting a proton from the aspartic acid. This process effectively splits the peptide into two different pieces while also regenerating the enzyme.(4)
Mechanism 1: Jaskólski et al. proposed a mechanism for aspartic protease whereby this process is one step.
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(Mechanism created using Chemdraw and information from Jaskólski et al.)
