Opening of a cryptic pocket is not suppressed by excluded volume effects of crowding

Cryptic pockets are transient structural features that could provide new opportunities for drug design. However, their utility would be greatly diminished if the probability of pocket opening were universally suppressed in cells due to the excluded volume effects of crowding. As a first step to addressing this concern, we study the effect of the synthetic crowder Ficoll 70, which has been shown to mimic the excluded volume effects of biomolecular crowding well, on the opening of a cryptic pocket in the interferon inhibitory domain (IID) of Ebola viral protein 35 (VP35). We find that this synthetic crowder has a negligible effect on the probability of cryptic pocket opening, as measured by both thiol labeling and hydrogen-deuterium exchange mass spectrometry (HDX-MS). The probability that a specific pocket opens may still be altered by the soft, enthalpic interactions of crowding, which will vary between different cellular environments. However, our results demonstrate that excluded volume effects that are at play in all crowded environments do not pose a universal barrier to targeting cryptic pockets.