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The desensitized channelrhodopsin-2 photointermediate contains 13 -cis, 15 -syn retinal Schiff base
(2021)
Channelrhodopsin-2 (ChR2) is a light-gated cation channel and was used to lay the foundations of optogenetics. Its dark state X-ray structure has been determined in 2017 for the wild-type, which is the prototype for all other ChR variants. However, the mechanistic understanding of the channel function is still incomplete in terms of structural changes after photon absorption by the retinal chromophore and in the framework of functional models. Hence, detailed information needs to be collected on the dark state as well as on the different photointermediates. For ChR2 detailed knowledge on the chromophore configuration in the different states is still missing and a consensus has not been achieved. Using DNP-enhanced solid-state MAS NMR spectroscopy on proteoliposome samples, we unambiguously determined the chromophore configuration in the desensitized state, and we show that this state occurs towards the end of the photocycle.
The excellent results of dispersion‐corrected density functional theory (DFT‐D) calculations for static systems have been well established over the past decade. The introduction of dynamics into DFT‐D calculations is a target, especially for the field of molecular NMR crystallography. Four 13C ss‐NMR calibration compounds are investigated by single‐crystal X‐ray diffraction, molecular dynamics and DFT‐D calculations. The crystal structure of 3‐methylglutaric acid is reported. The rotator phases of adamantane and hexamethylbenzene at room temperature are successfully reproduced in the molecular dynamics simulations. The calculated 13C chemical shifts of these compounds are in excellent agreement with experiment, with a root‐mean‐square deviation of 2.0 ppm. It is confirmed that a combination of classical molecular dynamics and DFT‐D chemical shift calculation improves the accuracy of calculated chemical shifts.