Pre-Gating Conformational Changes in the ChETA Variant of Channelrhodopsin-2 Monitored by Nanosecond IR Spectroscopy
| Autor: | Bernd-Joachim Schultz, Christian Bamann, Ernst Bamberg, Tom Resler, Víctor A. Lórenz-Fonfría, Ramona Schlesinger, Joachim Heberle |
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| Rok vydání: | 2015 |
| Předmět: |
Rhodopsin
Time Factors Photoisomerization Protein Conformation Static Electricity Infrared spectroscopy Photochemistry Vibration Biochemistry Catalysis chemistry.chemical_compound Colloid and Surface Chemistry Protein structure Spectroscopy Fourier Transform Infrared Side chain Spectroscopy Photolysis Chemistry Photodissociation Retinal General Chemistry Darkness Nanosecond Kinetics Mutation Retinaldehyde Diterpenes |
| Zdroj: | Journal of the American Chemical Society. 137:1850-1861 |
| ISSN: | 1520-5126 0002-7863 |
| DOI: | 10.1021/ja5108595 |
| Popis: | Light-gated ion permeation by channelrhodopsin-2 (ChR2) relies on the photoisomerization of the retinal chromophore and the subsequent photocycle, leading to the formation (on-gating) and decay (off-gating) of the conductive state. Here, we have analyzed the photocycle of a fast-cycling ChR2 variant (E123T mutation, also known as ChETA), by time-resolved UV/vis, step-scan FT-IR, and tunable quantum cascade laser IR spectroscopies with nanosecond resolution. Pre-gating conformational changes rise with a half-life of 200 ns, silent to UV/vis but detected by IR spectroscopy. They involve changes in the peptide backbone and in the H-bond of the side chain of the critical residue D156. Thus, the P1(500) intermediate must be separated into early and late states. Light-adapted ChR2 contains a mixture of all-trans and 13-cis retinal in a 70:30 ratio which are both photoactive. Analysis of ethylenic and fingerprint vibrations of retinal provides evidence that the 13-cis photocycle recovers in 1 ms. This recovery is faster than channel off-gating and most of the proton transfer reactions, implying that the 13-cis photocycle is of minor functional relevance for ChR2. |
| Databáze: | OpenAIRE |
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