HIV-1 protease flaps spontaneously close to the correct structure in simulations following manual placement of an inhibitor into the open state

Abstract

We report unrestrained, all-atom molecular dynamics simulations of HIV-1 protease (HIV-PR) with a continuum solvent model that reproducibly sample closing of the active site flaps following manual placement of a cyclic urea inhibitor into the substrate binding site of the open protease. The open form was obtained from the unbound, semi-open HIV-PR crystal structure, which we recently reported (Hornak, V.; et al. Proc. Natl. Acad. Sci. U.S.A. 2006, 103, 915-920.) to have spontaneously opened during unrestrained dynamics. In those simulations, the transiently open flaps always returned to the semi-open form that is observed in all crystal structures of the free protease. Here, we show that manual docking of the inhibitor reproducibly induces spontaneous conversion to the closed form as seen in all inhibitor-bound HIV-PR crystal structures. These simulations reproduced not only the greater degree of flap closure, but also the striking difference in flap "handedness" between bound and free enzyme. In most of the simulations, the final structures were highly accurate. Root-mean-square deviations (RMSD) from the crystal structure of the complex were approximately 1.5 A (averaged over the last 100 ps) for the inhibitor and each flap despite initial RMSD of 2-5 A for the inhibitors and 6-11 A for the flaps. Key hydrogen bonds were formed between the flap tips and between flaps and inhibitor that match those seen in the crystal structure. The results demonstrate that all-atom simulations have the ability to significantly improve poorly docked ligand conformations and reproduce large-scale receptor conformational changes that occur upon binding.

Figure 1

Crystal structures of HIV-PR: free (left, pdb 1HHP8) and with bound inhibitor (right, 1HVR7). A top view of the flaps is shown to illustrate the change in handedness that occurs upon inhibitor binding.

Figure 2

RMSD values during MD simulation following docking the inhibitor into fully open HIV-PR. The inhibitor (all atoms) and each flap (backbone) are compared to the bound, closed crystal structure of the complex, best-fit to the non-flaps portion of the protease. Final values fluctuate between 1–2 Å for the inhibitor and flap 1 and 1–3Å for flap2.

Figure 3

Simulated HIV-PR binding site and inhibitor after rough manual docking of the inhibitor into the open state (left) and after MD refinement (right). The crystallographic position of the inhibitor is shown in blue.