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Proc Natl Acad Sci U S A. 2015 Oct 27;112(43):E5796-804. doi: 10.1073/pnas.1511462112. Epub 2015 Oct 12.

Temporal evolution of helix hydration in a light-gated ion channel correlates with ion conductance.

Author information

1
Experimental Molecular Biophysics, Department of Physics, Freie Universität Berlin, 14195 Berlin, Germany; victor.lorenz@gmail.com joachim.heberle@fu-berlin.de.
2
Biophysical Chemistry, Max Planck Institute of Biophysics, 60438 Frankfurt am Main, Germany;
3
Experimental Molecular Biophysics, Department of Physics, Freie Universität Berlin, 14195 Berlin, Germany;
4
Genetic Biophysics, Department of Physics, Freie Universität Berlin, 14195 Berlin, Germany.

Abstract

The discovery of channelrhodopsins introduced a new class of light-gated ion channels, which when genetically encoded in host cells resulted in the development of optogenetics. Channelrhodopsin-2 from Chlamydomonas reinhardtii, CrChR2, is the most widely used optogenetic tool in neuroscience. To explore the connection between the gating mechanism and the influx and efflux of water molecules in CrChR2, we have integrated light-induced time-resolved infrared spectroscopy and electrophysiology. Cross-correlation analysis revealed that ion conductance tallies with peptide backbone amide I vibrational changes at 1,665(-) and 1,648(+) cm(-1). These two bands report on the hydration of transmembrane α-helices as concluded from vibrational coupling experiments. Lifetime distribution analysis shows that water influx proceeded in two temporally separated steps with time constants of 10 μs (30%) and 200 μs (70%), the latter phase concurrent with the start of ion conductance. Water efflux and the cessation of the ion conductance are synchronized as well, with a time constant of 10 ms. The temporal correlation between ion conductance and hydration of helices holds for fast (E123T) and slow (D156E) variants of CrChR2, strengthening its functional significance.

KEYWORDS:

channel gating; channelrhodopsin; infrared spectroscopy; optogenetics; time-resolved spectroscopy

PMID:
26460012
PMCID:
PMC4629336
DOI:
10.1073/pnas.1511462112
[Indexed for MEDLINE]
Free PMC Article

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