Viral BCL2 proteins (vBCL2s) help to sustain chronic infection of host proteins to inhibit apoptosis and autophagy. However, details of conformational changes in vBCL2s that enable binding to BH3Ds remain unknown. Using all-atom, multiple microsecond-long molecular dynamic simulations (totaling 17 μs) of the murine γ-herpesvirus 68 vBCL2 (M11), and statistical inference techniques, we show that regions of M11 transiently unfold and refold upon binding of the BH3D. Further, we show that this partial unfolding/refolding within M11 is mediated by a network of hydrophobic interactions, which includes residues that are 10 Å away from the BH3D binding cleft. We experimentally validate the role of these hydrophobic interactions by quantifying the impact of mutating these residues on binding to the Beclin1/BECN1 BH3D, demonstrating that these mutations adversely affect both protein stability and binding. To our knowledge, this is the first study detailing the binding-associated conformational changes and presence of long-range interactions within vBCL2s.
Anharmonicity in time-dependent conformational fluctuations is noted to be a key feature of functional dynamics of biomolecules. Although anharmonic events are rare, long-timescale (μs-ms and beyond) simulations facilitate probing of such events. We have previously developed quasi-anharmonic analysis to resolve higher-order spatial correlations and characterize anharmonicity in biomolecular simulations. In this article, we have extended this toolbox to resolve higher-order temporal correlations and built a scalable Python package called anharmonic conformational analysis (ANCA). ANCA has modules to: 1) measure anharmonicity in the form of higher-order statistics and its variation as a function of time, 2) output a storyboard representation of the simulations to identify key anharmonic conformational events, and 3) identify putative anharmonic conformational substates and visualization of transitions between these substates.
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