Abstract
Protein denaturation plays a crucial role in cellular processes. In this study, denaturation of HIV-1 Protease (PR) was investigated by all-atom MD simulations in explicit solvent. The PR dimer and monomer were simulated separately in 9 M acetic acid (9 M AcOH) solution and water to study the denaturation process of PR in acetic acid environment. Direct visualization of the denaturation dynamics that is readily available from such simulations has been presented. Our simulations in 9 M AcOH reveal that the PR denaturation begins by separation of dimer into intact monomers and it is only after this separation that the monomer units start denaturing. The denaturation of the monomers is flagged off by the loss of crucial interactions between the α-helix at C-terminal and surrounding β-strands. This causes the structure to transit from the equilibrium dynamics to random non-equilibrating dynamics. Residence time calculations indicate that denaturation occurs via direct interaction of the acetic acid molecules with certain regions of the protein in 9 M AcOH. All these observations have helped to decipher a picture of the early events in acetic acid denaturation of PR and have illustrated that the α-helix and the β-sheet at the C-terminus of a native and functional PR dimer should maintain both the stability and the function of the enzyme and thus present newer targets for blocking PR function.
Highlights
Protein folding/unfolding, dynamics and denaturation play crucial role in cellular processes and have been the subject of extensive investigation for the last several decades
In the simulation of the dimer in water, soon after 1 ns, the root mean square deviations (RMSD) fluctuates only between 0.25– 0.3 nm and the structure can be said to have quickly converged in the water simulation [Figure S1]
At around 10 ns during the acetic acid simulation, some sudden structural change in the PR conformation shifts it away from the native state and the PR fluctuates between different non-native states
Summary
Protein folding/unfolding, dynamics and denaturation play crucial role in cellular processes and have been the subject of extensive investigation for the last several decades. Using NMR investigations, acid denatured states [8] have been shown to be significantly different from the guanidine denatured tethered dimer of PR [9] or urea denatured PR precursor having the TFR extension at the N-terminal [10] All these experimental studies can only allude to the regions of the PR that are structured or denatured but cannot provide direct structural visualization of the dynamics of the protein as a whole in the native or denaturing environments. MD simulation has revealed that the unliganded protease predominantly populates the semi open conformation, with closed and fully open structures being a minor component of the overall ensemble and provides a model for such flap opening and closing [20] All these all-atom MD simulations of PR dealt with gross structure dynamics only and did not present a consolidated picture of the denaturation process, sequential loss of residue level interactions etc. For illustration we have chosen here acetic acid denaturation, but such studies can be extended to denaturation by other agents as well
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