June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
Critical role of central 139 loop in stability and binding selectivity of arrestin-1
Author Affiliations & Notes
  • Sergey Vishnivetskiy
    Pharmacology, Vanderbilt University, Nashville, TN
  • Faiza Baameur
    Pharmacology, Vanderbilt University, Nashville, TN
  • Kristen Findley
    Pharmacology, Vanderbilt University, Nashville, TN
  • Vsevolod Gurevich
    Pharmacology, Vanderbilt University, Nashville, TN
  • Footnotes
    Commercial Relationships Sergey Vishnivetskiy, None; Faiza Baameur, None; Kristen Findley, None; Vsevolod Gurevich, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 2458. doi:
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      Sergey Vishnivetskiy, Faiza Baameur, Kristen Findley, Vsevolod Gurevich; Critical role of central 139 loop in stability and binding selectivity of arrestin-1. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2458.

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

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Abstract

Purpose: To elucidate the functional role of central 139-loop on the receptor-binding surface of arrestin-1

Methods: Deletions of 139-loop and mutations that disrupt its interactions with other elements of the molecule were introduced in wild type arrestin-1 and its enhanced 3A mutant with high binding to unphosphorylated light-activated rhodopsin (Rh*). We tested binding selectivity and thermal stability of these mutants

Results: Using intra-molecular distance measurements by DEER we detected large movement of arrestin-1 139-loop to the side, away from the incoming receptor, upon its binding to active phosphorhodopsin (P-Rh*). Therefore, we hypothesized that elimination of this loop would promote rhodopsin binding. Deletions of increasing length in 139-loop progressively enhanced arrestin-1 binding to non-preferred forms of rhodopsin: Rh*, dark P-Rh, and phospho-opsin. The interactions of 139-loop with the neighboring finger loop involved in receptor binding were disrupted by charge eliminations and reversals on both sides. These milder perturbations increased arrestin-1 binding to non-preferred forms of rhodopsin, but to a lesser extent. In all cases an increase in binding to Rh*, dark P-Rh, and phospho-opsin correlated with a decrease in thermal stability of the mutants. Similar effects of deletions and mutations in 139-loop on both binding selectivity and thermal stability were observed in the context of WT arrestin-1 and an enhanced 3A mutant. which demonstrates higher binding to non-preferred forms of rhodopsin

Conclusions: Central 139 loop is an earlier unappreciated element stabilizing basal arrestin-1 conformation and precluding its binding to non-preferred forms of rhodopsin. Thermal stability of the basal state of arrestin-1 directly correlates with its selectivity for P-Rh*. NIH grants EY011500, GM077561, and GM081756 (VVG).

Keywords: 714 signal transduction • 659 protein structure/function • 648 photoreceptors  
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