INVOLVEMENT OF THE RECOVERIN C-TERMINAL SEGMENT IN RECOGNITION OF THE TARGET ENZYME RHODOPSIN KINASEстатья

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Дата последнего поиска статьи во внешних источниках: 18 июля 2013 г.

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1. Полный текст Involment_of_the_recoverin_C-treminal_segment_in_recognit... 820,7 КБ 19 мая 2012 [Ivan_Senin]

[1] Involvement of the recoverin c-terminal segment in recognition of the target enzyme rhodopsin kinase / E. Y. Zernii, K. E. Komolov, S. E. Permyakov et al. // Biochemical Journal. — 2011. — Vol. 435, no. 2. — P. 441–450. NCS (neuronal Ca2+ sensor) proteins belong to a family of calmodulin-related EF-hand Ca2+-binding proteins which, in spite of a high degree of structural similarity, are able to selectively recognize and regulate individual effector enzymes in a Ca2+-dependent manner. NCS proteins vary at their C-termini, which could therefore serve as structural control elements providing specific functions such as target recognition or Ca2+ sensitivity. Recoverin, an NCS protein operating in vision, regulates the activity of rhodopsin kinase, GRK1, in a Ca2+-dependent manner. In the present study, we investigated a series of recoverin forms that were mutated at the C-terminus. Using pull-down assays, surface plasmon resonance spectroscopy and rhodopsin phosphorylation assays, we demonstrated that truncation of recoverin at the C-terminus significantly reduced the affinity of recoverin for rhodopsin kinase. Site-directed mutagenesis of single amino acids in combination with structural analysis and computational modelling of the recoverin-kinase complex provided insight into the protein-protein interface between the kinase and the C-terminus of recoverin. Based on these results we suggest that Phe3 from the N-terminal helix of rhodopsin kinase and Lys192 from the C-terminal segment of recoverin form a cation-π interaction pair which is essential for target recognition by recoverin. Taken together, the results of the present study reveal a novel rhodopsin-kinase-binding site within the C-terminal region of recoverin, and highlights its significance for target recognition and regulation. [ DOI ]

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