Abstract
HIV-1 protease is an essential enzyme for viral particle maturation and is a target in the fight against HIV-1 infection worldwide. Several natural polymorphisms are also associated with drug resistance. Here, we utilized both pulsed electron double resonance, also called double electron-electron resonance, and NMR (15)N relaxation measurements to characterize equilibrium conformational sampling and backbone dynamics of an HIV-1 protease construct containing four specific natural polymorphisms commonly found in subtypes A, F, and CRF_01 A/E. Results show enhanced backbone dynamics, particularly in the flap region, and the persistence of a novel conformational ensemble that we hypothesize is an alternative flap orientation of a curled open state or an asymmetric configuration when interacting with inhibitors.
Highlights
HIV-1 protease is an essential enzyme for HIV maturation
We have shown that inhibitors vary in their ability to shift the conformational sampling to a closed state, with stronger protein-inhibitor interactions correlating with slower exchange dynamics having the greatest effect on flap closure for inactive HIV-1 PR [25]
Common Polymorphisms May Be Responsible for the Increased Backbone Dynamics and Altered Conformational Sampling—As shown here and in our previous work [14, 25], both PR5 and CRF_01 A/E have a 10 –20% fractional occupancy of the hypothesized curled open flap conformation and have increased backbone dynamics compared with subtype B
Summary
HIV-1 protease is an essential enzyme for HIV maturation. Results: Select and naturally occurring polymorphisms alter the conformational sampling and backbone dynamics of HIV-1 protease. Our previous studies have shown that this method is successful in characterizing ligand-induced shifts in the conformational ensembles [23, 26], that natural polymorphisms and drug pressure-selected mutations alter the fractional occupancy of the conformational ensemble [14, 24, 25, 27], and that there are correlations of fractional occupancy with enzyme kinetics and inhibitor Ki values [25,26,27] Taken together, these findings indicate that changes in dynamics and conformational sampling likely play important roles in lowering inhibitor effects seen with drug pressure-selected mutations. We hypothesize that the removal of this salt bridge is likely responsible for destabilizing the compact fold of the flap, inducing an alternative flap conformation that may be a curled open or asymmetric conformation; concurrently removal of this salt bridge interaction leads to increased backbone dynamics
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