April 2014
Volume 55, Issue 13
ARVO Annual Meeting Abstract  |   April 2014
Rhodopsin binding-induced conformational changes in arrestin-1
Author Affiliations & Notes
  • Sergey A Vishnivetskiy
    Pharmacology, Vanderbilt University, Nashville, TN
  • Qiuyan Chen
    Pharmacology, Vanderbilt University, Nashville, TN
  • Miyeon Kim
    UCLA, Los Angeles, CA
  • Ned Van Eps
    UCLA, Los Angeles, CA
  • Tiandi Zhuang
    Pharmacology, Vanderbilt University, Nashville, TN
  • Wayne L Hubbell
    UCLA, Los Angeles, CA
  • Charles R Sanders
    Pharmacology, Vanderbilt University, Nashville, TN
  • Eric Xu
    Van Andel Institute, Grand Rapids, MI
  • Vsevolod V Gurevich
    Pharmacology, Vanderbilt University, Nashville, TN
  • Footnotes
    Commercial Relationships Sergey Vishnivetskiy, None; Qiuyan Chen, None; Miyeon Kim, None; Ned Van Eps, None; Tiandi Zhuang, None; Wayne Hubbell, None; Charles Sanders, None; Eric Xu, None; Vsevolod Gurevich, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 3028. doi:
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      Sergey A Vishnivetskiy, Qiuyan Chen, Miyeon Kim, Ned Van Eps, Tiandi Zhuang, Wayne L Hubbell, Charles R Sanders, Eric Xu, Vsevolod V Gurevich; Rhodopsin binding-induced conformational changes in arrestin-1. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3028.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: Arrestins specifically bind active phosphorylated GPCRs, block their coupling to G proteins, and redirect signaling to alternative pathways. Several lines of evidence suggest that the conformation of receptor-bound arrestin is different from the free arrestin in the cytoplasm.

Methods: We explored the conformation using three different methods. Intra-molecular distances in arrestin-1, which was either free or bound to phosphorylated light-activated rhodopsin, were measured with the pulse EPR technique DEER. Rhodopsin in native disc membranes and reconstituted in monomeric form into HDL particles was used. Structural changes in constitutively monomeric arrestin-1 labeled with 13C and 15N induced by its binding to different functional forms of unlabeled rhodopsin in bicelles were detected using NMR. The structure of the complex of enhanced phosphorylation-independent arrestin-1 with rhodopsin carrying activating mutations was determined by X-ray crystallography.

Results: All three methods identified an extensive receptor-binding surface, revealed significant receptor-induced rearrangements, and showed that receptor-bound arrestin does not assume a rigid “active” conformation, but remains unexpectedly flexible. This flexibility likely explains how receptor-bound non-visual arrestins can initiate many different signaling cascades interacting with numerous proteins.

Conclusions: The flexibility of the receptor-bound arrestin likely explains why different methods reveal distinct receptor-induced conformational changes in arrestin-1. Whereas the binding to light-activated unphosphorylated and to phosphorylated inactive rhodopsin induces only local structural perturbations, the binding to the high-affinity targets, active phospho-rhodopsin and phospho-opsin, globally changes the structure and makes it a lot more flexible. Thus, receptor-bound arrestin explores numerous conformational states, which apparently allows it to fit an amazing variety of binding partners. Supported by NIH grant EY011500.

Keywords: 648 photoreceptors • 658 protein purification and characterization • 659 protein structure/function  

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