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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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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.
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.
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.
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.
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