Figure 7.

IR difference spectra in the carboxylic region of wild-type ChR2 (top panel) and the D156E variant (bottom panel). For the wild-type, spectra at 6 ms in H₂O and at 18 ms in D₂O after the exciting laser pulse were extracted from the QCL data. The spectrum for the D156E variant was recorded under continuous illumination at 450 nm, representing mostly the P₃⁵²⁰ state. The negative band was assigned to the C=O stretching vibration of protonated D156 (top panel) and E156 (bottom panel), respectively. Vibrational isotope effect leads to a spectral upshift of 2 cm⁻¹ in wild-type ChR2 (top panel). The D156E variant exhibits a vibrational downshift of 8 cm⁻¹ in D₂O, typical for H/D exchange (bottom panel). To see this figure in color, go online.