Single point mutation induced alterations in the equilibrium structural transitions on the folding landscape of HIV-1 protease.

Equilibrium folding-unfolding transitions are hard to study in HIV-1 protease (PR) because of its autolytic properties. Further, the protease exhibits many tolerant point mutations some of which also impart drug resistance to the protein. It is conceivable that the mutations affect protein's function by altering its folding characteristics; these would clearly depend on the nature of the mutations themselves. In this background, we report here NMR studies on the effects of D25 N mutation, which removes one negative charge from the protein at the active site, on the equilibrium folding behaviour of PR starting from its acetic acid denatured state. It is observed that in PRD25N two slowly exchanging conformations are present at the N-terminal. One of them is similar to that of PR. Though the conformational and dynamics preferences of PR and PRD25N are fairly similar in 9 M acetic acid, they seem to undergo different folding transitions when acetic acid concentration is reduced. The differences are seen in the active site, in the flap, and in the hinge of the flap regions. The present study suggests that such differences, though different in detail, would occur for other mutations as well, and also for different initial denatured states. These would have significant regulatory implications for the efficacy of protease function.