Channelrhodopsin is a light-gated cation channel whose reaction cycle involves proton-transfer reactions. Understanding how channelrhodopsin works is important, as it may assist in designing channelrhodopsin variants with specific properties for optogenetics applications.To dissect structural elements that may act as gates and to explore how protein and water dynamics respond to changes in the protonation state, we combined extensive bioinformatics analyses with molecular dynamics simulations of channelrhodopsin and of bacteriorhodopsin mutants that model specific channelrhodopsin interactions, or have altered proton-transfer kinetics. In some of these mutants, we find that perturbation of specific hydrogen bonds is coupled to the rapid formation of water bridges that could assist proton transfer. In simulations on channelrhodopsin embedded in hydrated lipid membranes, the dynamics of water wires and hydrogen bonds inter-connecting remote regions of the protein are tightly coupled to the protonation state.This work has been supported by the Marie Curie International Reintegration Award FP7-PEOPLE-2010-RG 276920 and by the SFB 1078 ‘Protonation Dynamics in Protein Function'.