Fernández Sampedro, Miguel A.Invergo, Brandon M., 1982-Ramon, EvaBertranpetit, Jaume, 1952-Garriga, Pere2016-03-292016-03-292016Fernández-Sampedro MA, Invergo BM, Ramon E, Bertranpetit J, Garriga P. Functional role of positively selected amino acid substitutions in mammalian rhodopsin evolution. Scientific reports. 2016; 6: 21570. DOI 10.1038/srep215702045-2322http://hdl.handle.net/10230/26029Visual rhodopsins are membrane proteins that function as light photoreceptors in the vertebrate retina. Specific amino acids have been positively selected in visual pigments during mammal evolution, which, as products of adaptive selection, would be at the base of important functional innovations. We have analyzed the top candidates for positive selection at the specific amino acids and the corresponding reverse changes (F13M, Q225R and A346S) in order to unravel the structural and functional consequences of these important sites in rhodopsin evolution. We have constructed, expressed and immunopurified the corresponding mutated pigments and analyzed their molecular phenotypes. We find that position 13 is very important for the folding of the receptor and also for proper protein glycosylation. Position 225 appears to be important for the function of the protein affecting the G-protein activation process, and position 346 would also regulate functionality of the receptor by enhancing G-protein activation and presumably affecting protein phosphorylation by rhodopsin kinase. Our results represent a link between the evolutionary analysis, which pinpoints the specific amino acid positions in the adaptive process, and the structural and functional analysis, closer to the phenotype, making biochemical sense of specific selected genetic sequences in rhodopsin evolution.application/pdfeng© Nature Publishing Group. http://www.nature.com/articles/srep21570/nThis work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/Aminoàcidsinfo:eu-repo/semantics/articlehttp://dx.doi.org/10.1038/srep21570info:eu-repo/semantics/openAccess