Drug-resistance-associated mutation in essential proteins of the viral life cycle is a major concern in anti-retroviral therapy. M46I, a non-active site mutation in HIV-1 protease has been clinically associated with saquinavir resistance in HIV patients. A 100 ns molecular dynamics (MD) simulation and MM-PBSA calculations were performed to study the molecular mechanism of M46I-mutation-based saquinavir resistance. In order to acquire deeper insight into the drug-resistance mechanism, the flap curling, closed/semi-open/open conformations, and active site compactness were studied. The M46I mutation significantly affects the energetics and conformational stability of HIV-1 protease in terms of RMSD, RMSF, Rg, SASA, and hydrogen formation potential. This mutation significantly decreased van der Waals interaction and binding free energy (∆G) in the M46I–saquinavir complex and induced inward flap curling and a wider opening of the flaps for most of the MD simulation period. The predominant open conformation was reduced, but inward flap curling/active site compactness was increased in the presence of saquinavir in M46I HIV-1 protease. In conclusion, the M46I mutation induced structural dynamics changes that weaken the protease grip on saquinavir without distorting the active site of the protein. The produced information may be utilized for the discovery of inhibitor(s) against drug-resistant HIV-1 protease.