AbstractProteins are known to be dynamic molecules that undergo conformational fluctuations. A fundamental issue that remains to be clarified is whether there is a linkage between the dynamic nature of proteins and their catalytic function. Of particular interest are those conformations accessible near global free energy minima which are in equilibrium and separated by low energy barriers, also called substates. Because proteins only function in their native state, interconversions between these substates are important. As a result, a complete understanding of the mechanisms governing the interconversions between these substates not only sheds light on how the enzyme works, but also has profound and practical implications for revealing new approaches to drug design.In this work, both experimental and theoretical tools have been employed collaboratively to explore the structural and dynamic features of HIV-1 protease, aimed to obtain a better understanding of conformational transitions of the enzyme, which may open new avenues in the design of more effective treatment regimes. Here, we present a hypothesis, based on microsecond molecular dynamics simulations of an apo HIV-1 protease with explicit solvent, describing how the twisting of the backbone of the flap tips transforms the geometry of the β-hairpin structure of each flap from the `closed' conformation to the `semi-open' one, most likely owing to the intrinsic flexibility of the glycine residues. In addition, we suggest that it is the various binding interactions within the protease dimer interface that govern the gating properties of the flaps; the opening of the flaps most likely results from the concerted partial dissociation of the dimer interface facilitated by water dynamics. Moreover, to explore how resistance caused by protease mutations arises, we collaborated with EPR experimentalists and performed a series of MD simulations on the spin-labeled wild-type and multi-drug resistant proteases. The combined analysis suggests that the semi-open form is most likely the dominant configuration; mutations conferring drug resistance may alter either the conformation of the flaps or the mobility of the flaps, or both.