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J Biol Chem. 2019 Aug 9. pii: jbc.RA119.010089. doi: 10.1074/jbc.RA119.010089. [Epub ahead of print]

Cryo-EM structure of the native rhodopsin dimer in nanodiscs.

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University of Toronto, Canada.
Max Planck Institute of Biochemistry, Germany.
Case Western Reserve University.
Gavin Herbert Eye Institute, University of California, Irvine, United States.
Faculty of Chemistry, Biological and Chemical Research Centre, University of Warsaw, Poland.
Biological and Chemical Research Centre; University of Warsaw, Poland.
Structural and Computational Biology Unit, European Molecular Biology Laboratory, Germany.
Molecular Structural Biology, Max Planck Institute of Biochemistry, Germany.
Max Planck Institute of Biochemistry.
Biochemistry, University of Toronto, Canada.
Department of Ophthalmology, Gavin Herbert Eye Institute, University of California, Irvine, United States.


Imaging of rod photoreceptor outer-segment disc membranes by atomic force microscopy (AFM) and cryo-electron tomography has revealed that the visual pigment rhodopsin, a prototypical class A G protein-coupled receptor (GPCR), can organize as rows of dimers. GPCR dimerization and oligomerization offer possibilities for allosteric regulation of GPCR activity, but the detailed structures and mechanism remain elusive. In this investigation, we made use of the high rhodopsin density in the native disc membranes and of a bifunctional cross-linker that preserves the native rhodopsin arrangement by covalently tethering rhodopsins via Lys residue side chains. We purified cross-linked rhodopsin dimers and reconstituted them into nanodiscs for cryo-EM analysis. We present cryo-EM structures of the crosslinked rhodopsin dimer as well as a rhodopsin dimer reconstituted into nanodiscs from purified monomers. We demonstrate the presence of a preferential two-fold symmetrical dimerization interface mediated by transmembrane helix 1 and the cytoplasmic helix 8 of rhodopsin. We confirmed this dimer interface by double electron-electron resonance (DEER) measurements of spin-labeled rhodopsin. We propose that this interface and the arrangement of two protomers is a prerequisite for the formation of the observed rows of dimers. We anticipate that the approach outlined here could be extended to other GPCRs or membrane receptors to better understand specific receptor dimerization mechanisms.


G protein-coupled receptor (GPCR); receptor; retina; retinoid-binding protein; rhodopsin

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