Proton transfer reactions in the red light-activatable channelrhodopsin variant ReaChR and their relevance for its function
| Autor: | Peter Hegemann, Eglof Ritter, Christiane Grimm, Benjamin S. Krause, Joel C.D. Kaufmann, Franz Bartl |
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| Rok vydání: | 2017 |
| Předmět: |
Models
Molecular 0301 basic medicine Rhodopsin Light Proton Protein Conformation Stereochemistry Recombinant Fusion Proteins Channelrhodopsin Chlamydomonas reinhardtii Molecular Dynamics Simulation Photochemistry Biochemistry 03 medical and health sciences chemistry.chemical_compound Protein structure Chlorophyta Catalytic Domain Spectroscopy Fourier Transform Infrared Humans Protein Isoforms Molecular Biology Plant Proteins Schiff base 030102 biochemistry & molecular biology biology Protein Stability Hydrogen bond Algal Proteins Hydrogen Bonding Cell Biology biology.organism_classification Recombinant Proteins Electrophysiological Phenomena HEK293 Cells 030104 developmental biology Amino Acid Substitution chemistry Mutation Mutagenesis Site-Directed biology.protein Selectivity Molecular Biophysics |
| Zdroj: | Journal of Biological Chemistry. 292:14205-14216 |
| ISSN: | 0021-9258 |
| Popis: | Channelrhodopsins (ChRs) are light-gated ion channels widely used for activating selected cells in large cellular networks. ChR variants with a red-shifted absorption maximum, such as the modified Volvox carteri ChR1 red-activatable channelrhodopsin (“ReaChR,” λmax = 527 nm), are of particular interest because longer wavelengths allow optical excitation of cells in deeper layers of organic tissue. In all ChRs investigated so far, proton transfer reactions and hydrogen bond changes are crucial for the formation of the ion-conducting pore and the selectivity for protons versus cations, such as Na+, K+, and Ca2+ (1). By using a combination of electrophysiological measurements and UV-visible and FTIR spectroscopy, we characterized the proton transfer events in the photocycle of ReaChR and describe their relevance for its function. 1) The central gate residue Glu130 (Glu90 in Chlamydomonas reinhardtii (Cr) ChR2) (i) undergoes a hydrogen bond change in D → K transition and (ii) deprotonates in K → M transition. Its negative charge in the open state is decisive for proton selectivity. 2) The counter-ion Asp293 (Asp253 in CrChR2) receives the retinal Schiff base proton during M-state formation. Starting from M, a photocycle branching occurs involving (i) a direct M → D transition and (ii) formation of late photointermediates N and O. 3) The DC pair residue Asp196 (Asp156 in CrChR2) deprotonates in N → O transition. Interestingly, the D196N mutation increases 15-syn-retinal at the expense of 15-anti, which is the predominant isomer in the wild type, and abolishes the peak current in electrophysiological measurements. This suggests that the peak current is formed by 15-anti species, whereas 15-syn species contribute only to the stationary current. |
| Databáze: | OpenAIRE |
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