Arrestin with a Single Amino Acid Substitution Quenches Light-Activated Rhodopsin in a Phosphorylation-Independent Fashion

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• 作者：Mark P. Gray-Keller ; Peter B. Detwiler ; Jeffrey L. Benovic ; and Vsevolod V. Gurevich
• 刊名：Biochemistry
• 出版年：1997
• 出版时间：June 10, 1997
• 年：1997
• 卷：36
• 期：23
• 页码：7058 - 7063
• 全文大小：117K
• 年卷期：v.36,no.23(June 10, 1997)
• ISSN：1520-4995

文摘

Arrestins are members of a superfamily of regulatory proteins thatparticipate in the terminationof G protein-mediated signal transduction. In thephototransduction cascade of vertebrate rods, whichserves as a prototypical G protein-mediated signaling pathway, thebinding of visual arrestin is stimulatedby phosphorylation of the C-terminus of photoactivated rhodopsin (Rh*).Arrestin is very selective towardlight-activated phosphorhodopsin (P-Rh*). Previously we reportedthat a single amino acid substitutionin arrestin, Arg175Gln, results in a dramatic increase in arrestinbinding to Rh* [Gurevich, V. V., &Benovic, J. L. (1995) J. Biol. Chem. 270,6010-6016]. Here we demonstrate that a similarmutant,arrestin(R175E), binds to light-activated rhodopsin independent ofphosphorylation. Arrestin(R175E) bindswith high affinity not only to P-Rh* and Rh* but also tolight-activated truncated rhodopsin in which theC-terminus phosphorylation sites have been proteolytically removed.In an in vitro assay that monitoredrhodopsin-dependent activation of cGMP phosphodiesterase (PDE), wildtype arrestin quenched PDEresponse only when ATP was present to support rhodopsinphosphorylation. In contrast, as little as 30nM arrestin(R175E) effectively quenched PDE activation in theabsence of ATP. Arrestin(R175E) hadno effect when the lifetime of Rh* no longer contributed to the timecourse of PDE activity, suggestingthat it disrupts signal transduction at the level ofrhodopsin-transducin interaction.