3.E.1.7.1
Channelrhodopsin-1 (chlamyrhodopsin-3) (ChR1; Cop3; CSOA) (light-gated cation (H⁺, Na⁺, K⁺, and Ca²⁺) channel) (Nagel et al., 2003). TMSs 1 and 2 are the main structures involved in desensitization involving the stabilization of the protein's conformation and the alteration of the charge distribution around the retinal-Schiff base (Zamani et al. 2017). Replacing the glutamate located at the central gate of the ion channel with positively charged amino acyl residues reverses the ion selectivity and allows anion (chloride, Cl^-) conduction (Zhang et al. 2019).  zlight-gated channelrhodopsin sparks proton-induced calcium release in guard cells (Huang et al. 2023).  The mechanisms of cation transport and valence selectivity through the channelrhodopsin chimera, C1C2 (ChR1/ChR2), in the high- and low-conducting open states have been examined (Prignano et al. 2024). Electrophysiology measurements identified a single-residue substitution within the central gate, N297D, that increased Ca²⁺ permeability vs. Na⁺ by nearly two-fold at peak current, but less so at stationary current. Molecular models of dimeric wild-type C1C2 and N297D mutant channels were examined in both open states and the PMF profiles for Na⁺ and Ca²⁺ permeation through each protein using well-tempered/multiple-walker metadynamics were determined. Results of these studies agree well with experimental measurements and demonstrated that the pore entrance on the extracellular side differs from original predictions and is actually located in a gap between helices I and II. Cation transport occurs via a relay mechanism where cations are passed between flexible carboxylate sidechains lining the full length of the pore by side chain swinging, like a monkey swinging on vines. In the mutant channel, residue D297 enhances Ca²⁺ permeability by mediating the handoff between the central and cytosolic binding sites via direct coordination and side chain swinging. Prignano et al. 2024 also found that altered cation binding affinities at both the extracellular entrance and the central binding sites underlie the distinct transport properties of the low-conducting open state. This facilitates an understanding of ion selectivity and permeation in cation channelrhodopsins.