title: Light-dark adaptation of channelrhodopsin C128T mutant.
abstract: Channelrhodopsins are microbial type rhodopsins that operate as light-gated ion channels. Largely prolonged lifetimes of the conducting state of channelrhodopsin-2 may be achieved by mutations of crucial single amino acids, i.e. cysteine 128. Such mutants are of great scientific interest in the field of neurophysiology because they allow neurons to be switched on and off on demand (step function rhodopsins). Due to their slow photocycle, structural alterations of these proteins can be studied by vibrational spectroscopy in more detail than possible with wild type. Here, we present spectroscopic evidence that the photocycle of the C128T mutant involves three different dark-adapted states that are populated according to the wavelength and duration of the preceding illumination. Our results suggest an important role of multiphoton reactions and the previously described side reaction for dark state regeneration. Structural changes that cause formation and depletion of the assumed ion conducting state P520 are only small and follow larger changes that occur early and late in the photocycle, respectively. They require only minor structural rearrangements of amino acids near the retinal binding pocket and are triggered by all-trans/13-cis retinal isomerization, although additional isomerizations are also involved in the photocycle. We will discuss an extended photocycle model of this mutant on the basis of spectroscopic and electrophysiological data.
authors: Institut für Medizinische Physik und Biophysik, Charité-Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany. eglof.ritter@charite.de
authors: Eglof
authors: E
authors: Ritter
authors: Patrick
authors: P
authors: Piwowarski
authors: Peter
authors: P
authors: Hegemann
authors: Franz J
authors: FJ
authors: Bartl
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002522:Chlorocebus aethiops; D003623:Dark Adaptation / Q000502:physiology*; D020125:Mutation, Missense; D012172:Retinaldehyde / Q000235:genetics / Q000378:metabolism*; D012243:Rhodopsin / Q000235:genetics / Q000378:metabolism*
chemical_list: D012243:Rhodopsin; D012172:Retinaldehyde