Abstract
Proteorhodopsin (PR) is a microbial proton pump that is ubiquitous in marine environments and may play an important role in the oceanic carbon cycle. Photoisomerization of the retinal chromophore in PR leads to a series of proton transfers between specific acidic amino acid residues and the Schiff base of retinal, culminating in a proton motive force to facilitate ATP synthesis. The proton donor in a similar retinal protein, bacteriorhodopsin, acts as a latch to allow the influx of bulk water. However, it is unclear if the proton donor in PR, E108, utilizes the same latch mechanism to become internally hydrated. Here, we used molecular dynamics simulations to model the changes in internal hydration of the blue variant of PR during photoactivation with the proton donor in protonated and deprotonated states. We find that there is a stark contrast in the levels of internal hydration of the cytoplasmic half of PR based on the protonation state of E108. Instead of a latch mechanism, deprotonation of E108 acts as a gate, taking advantage of a nearby polar residue (S61) to promote the formation of a stable water wire from bulk cytoplasm to the retinal-binding pocket over hundreds of nanoseconds. No large-scale conformational changes occur in PR over the microsecond timescale. This subtle yet clear difference in the effect of deprotonation of the proton donor in PR may help explain why the photointermediates that involve the proton donor (i.e., M and N states) have timescales that are orders of magnitude different from the archaeal proton pump, bacteriorhodopsin. In general, our study highlights the importance of understanding how structural fluctuations lead to differences in the way that retinal proteins accomplish the same task.