Fig. 2: Engineering of Calcium-permeable ChannelRhodopsins (CapChRs).
From: Calcium-permeable channelrhodopsins for the photocontrol of calcium signalling
A Design strategy for CapChR1 (green) and CapChR2 (orange). Pore helices coloured in dark grey and miscellaneous helices in light grey. B and C Exemplary CapChR2 homology model depicting mutated residues and their intended effects (based on structure data, PDB ID: 6EID, https://doi.org/10.2210/pdb6EID/pdb). Grey mesh represents water accessibility according to MD simulations. D and H Representative photocurrent traces of CapChR1 (green) and CapChR2 (orange) respectively, recorded from −80 to +40 mV in 20 mV steps in ND7/23 cells (blue bar: illumination with saturating, 470 nm light; ~1.9 mW/mm²). E and I Current–voltage relationships for CapChR1 and CapChR2 under varying extracellular conditions (shadows represent the SEM; normalized to high [NaCl]e at −80 mV). F and J Stationary photocurrents for CapChR1 and CapChR2 at the designated extracellular conditions (mean ± SEM). G and K Estimated reversal potential (Erev) for CapChR1 and CapChR2 at the designated ionic conditions (Box middle line: Mean; Box outer edges ± SEM; Box whiskers: 1.5 × SEM). Number of replicates in E [NaCl]e: n = 14; [CaCl2]e: n = 12; [NMGCl]e: n = 6, in F) [NaCl]e: n = 14; [CaCl2]e pH 7.2: n = 12; [CaCl2]e pH 9: n = 5; [NMGCl]e: n = 5; [KCl]e: n = 3; [MgCl2]e: n = 4, in G) ([NaCl]e: n = 14; [CaCl2]e pH 7.2: n = 12; [CaCl2]e pH 9: n = 5; [NMGCl]e: n = 6; [KCl]e: n = 3; [MgCl2]e: n = 4), in I) ([NaCl]e: n = 10; [CaCl2]e: n = 7; [NMGCl]e: n = 6), in J [NaCl]e: n = 10; [CaCl2]e pH 7.2: n = 7; [CaCl2]e pH 9: n = 5; [NMGCl]e: n = 6; [KCl]e: n = 3; [MgCl2]e: n = 3, in K) [NaCl]e: n = 10; [CaCl2]e pH 7.2: n = 7; [CaCl2]e pH 9: n = 5; [NMGCl]e: n = 6; [KCl]e: n = 3; [MgCl2]e: n = 3. n = X biologically independent cells. EC extracellular, IC intracellular.
