The new and vibrant field of optogenetics was founded by the seminal discovery
of channelrhodopsin, the first light-gated cation channel. Despite the
numerous applications that have revolutionised neurophysiology, the functional
mechanism is far from understood on the molecular level. An arsenal of
biophysical techniques has been established in the last decades of research on
microbial rhodopsins. However, application of these techniques is hampered by
the duration and the complexity of the photoreaction of channelrhodopsin
compared with other microbial rhodopsins. A particular interest in resolving
the molecular mechanism lies in the structural changes that lead to channel
opening and closure. Here, we review the current structural and mechanistic
knowledge that has been accomplished by integrating the static structure
provided by X-ray crystallography and electron microscopy with time-resolved
spectroscopic and electrophysiological techniques. The dynamical reactions of
the chromophore are effectively coupled to structural changes of the protein,
as shown by ultrafast spectroscopy. The hierarchical sequence of structural
changes in the protein backbone that spans the time range from 10− 12 s to 10−
3 s prepares the channel to open and, consequently, cations can pass. Proton
transfer reactions that are associated with channel gating have been resolved.
In particular, glutamate 253 and aspartic acid 156 were identified as proton
acceptor and donor to the retinal Schiff base. The reprotonation of the latter
is the critical determinant for channel closure. The proton pathway that
eventually leads to proton pumping is also discussed
