Conformational equilibria of light-activated rhodopsin in nanodiscs

Ned Van Eps, Lydia N. Caro, Takefumi Morizumi, Ana Karin Kusnetzow, Michal Szczepek, Klaus Peter Hofmann, Timothy H. Bayburt, Stephen Sligar, Oliver P. Ernst, Wayne L. Hubbell

Research output: Contribution to journalArticle

Abstract

Conformational equilibria of G-protein-coupled receptors (GPCRs) are intimately involved in intracellular signaling. Here conformational substates of the GPCR rhodopsin are investigated in micelles of dodecyl maltoside (DDM) and in phospholipid nanodiscs by monitoring the spatial positions of transmembrane helices 6 and 7 at the cytoplasmic surface using site-directed spin labeling and double electron-electron resonance spectroscopy. The photoactivated receptor in DDM is dominated by one conformation with weak pH dependence. In nanodiscs, however, an ensemble of pH-dependent conformational substates is observed, even at pH 6.0 where the MIIbH+ form defined by proton uptake and optical spectroscopic methods is reported to be the sole species present in native disk membranes. In nanodiscs, the ensemble of substates in the photoactivated receptor spontaneously decays to that characteristic of the inactive state with a lifetime of ∼16 min at 20 °C. Importantly, transducin binding to the activated receptor selects a subset of the ensemble in which multiple substates are apparently retained. The results indicate that in a native-like lipid environment rhodopsin activation is not analogous to a simple binary switch between two defined conformations, but the activated receptor is in equilibrium between multiple conformers that in principle could recognize different binding partners.

Original languageEnglish (US)
Pages (from-to)E3268-E3275
JournalProceedings of the National Academy of Sciences of the United States of America
Volume114
Issue number16
DOIs
StatePublished - Apr 18 2017

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Keywords

  • Conformational heterogeneity
  • Double electron-electron resonance
  • GPCR
  • Nanodiscs
  • Rhodopsin

ASJC Scopus subject areas

  • General

Cite this

Van Eps, N., Caro, L. N., Morizumi, T., Kusnetzow, A. K., Szczepek, M., Hofmann, K. P., Bayburt, T. H., Sligar, S., Ernst, O. P., & Hubbell, W. L. (2017). Conformational equilibria of light-activated rhodopsin in nanodiscs. Proceedings of the National Academy of Sciences of the United States of America, 114(16), E3268-E3275. https://doi.org/10.1073/pnas.1620405114