STRUCTURAL AND FUNCTIONAL-STUDIES IN-VITRO ON THE P6 PROTEIN FROM THEHIV-1 GAG OPEN READING FRAME

Protein p6 from HIV-1 gag open reading frame is reported to affect bot h the final phase of assembly of the viral particle and the early stag e of the gag polyprotein maturation in vitro. Two separate hypotheses have been proposed, on only one of these reported effects. We think th at both observations may be eventually explained if p6 protein strongl y inhibits the HIV-1 proteinase. Protein p6 was synthesised by solid-p hase peptide synthesis. Several methods of folding the p6 protein were tested, each resulting in the random structure according to both CD a nd 1D proton NMR spectra. A uniformly high exposure of NH protons to t he solution was confirmed by temperature-dependent NMR spectra and iso tope exchange experiments. Thus the p6 protein does not have any rigid conformation in solution. A rigid structure is not formed after furth er cleavage by HIV-1 proteinase as neither the protein nor its fragmen ts are cleaved by this proteinase. In addition, the p6 protein itself does not act as inhibitor of HIV-1 proteinase. This excludes a direct role of p6 protein and supports the hypothesis that p6 is involved in forming the appropriate structure of gag polyprotein precursor. The ro le of slowly cleaved tight gag-proteinase in the final stage of matura tion may be to slow down maturation of the precursor polyproteins prio r to their transport to final location in the membrane.