May 2004
Volume 45, Issue 13
ARVO Annual Meeting Abstract  |   May 2004
Very Few Arrestin Residues Determine Its Receptor Preference.
Author Affiliations & Notes
  • S.A. Vishnivetskiy
    Pharmacology, Vanderbilt Univesity, Nashville, TN
  • J.L. Benovic
    Kimmel Cancer Institute, Thomas Jefferson Univesity, Philadelphia, PA
  • V.V. Gurevich
    Pharmacology, Vanderbilt Univesity, Nashville, TN
  • Footnotes
    Commercial Relationships  S.A. Vishnivetskiy, None; J.L. Benovic, None; V.V. Gurevich, None.
  • Footnotes
    Support  NIH Grants EY11500, GM63097 (VVG), GM44944, GM 47417 (JLB).
Investigative Ophthalmology & Visual Science May 2004, Vol.45, 1270. doi:
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      S.A. Vishnivetskiy, J.L. Benovic, V.V. Gurevich; Very Few Arrestin Residues Determine Its Receptor Preference. . Invest. Ophthalmol. Vis. Sci. 2004;45(13):1270.

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

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Abstract: : Purpose: To elucidate the amino acid residues responsible for the preferential binding of different arrestin proteins to particular receptors. Methods: We took advantage of a dramatic difference between visual arrestin and arrestin2 binding to phosphorylated light–activated rhodopsin (P–Rh*) and the phosphorylated carbachol–activated m2 muscarinic cholinergic receptor(P–m2 mAChR*). We constructed over 20 visual arrestin–based mutants containing from one to five arrestin2 residues, expressed them in a cell–free translation system in radiolabeled form, and tested their binding to both receptors in vitro. Results: We have recently identified two arrestin elements that dictate its receptor specificity. Residues 49–90 and 237–268 in visual arrestin and the homologous regions in arrestin2 are localized on the concave sides of the two arrestin domains and are largely responsible for their receptor preference. In these two elements only 35 amino acids are different, 22 of which are non–conservative substitutions. These residues in visual arrestin were mutated singly or in groups into their arrestin2 counterparts. The binding of the visual arrestin–based mutants to P–Rh* and P–m2 mAChR* was compared to that of the parental proteins and chimeras containing whole N– or C–domain regions from arrestin2. Several point mutations and their combinations reduce visual arrestin binding to P–Rh* and increase its binding to P–m2 mAChR* at the same time. We interpreted these changes as partial shift in receptor preference towards that of arrestin2. In particular, mutations L51Y, V52L, G54E, K55R and V90S in the N–domain and K257C, Q265D and K267T in the C–domain are the most effective in changing receptor specificity. The majority of these residues are exposed in the cavity of the respective domains. However, Val–90 is buried in the beta–strand sandwich and appears to enhance the rigidity of the N–domain in the highly specialized visual arrestin via multiple intra–molecular interactions. Conclusions: A limited number of residues in each arrestin domain likely determine its preference for the cognate receptor.

Keywords: signal transduction • protein structure/function • photoreceptors 

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