International Research Journal of Biological Sciences ___________________________________ ISSN 2278-3202Vol. 2(7), 6-11, July (2013) Int. Res. J. Biological Sci. International Science Congress Association 6 Evolutionary Analysis and Motif Discovery in Rhodopsin from VertebratesKumar Akash and Dwivedi Vivek Dhar* Department of Bioinformatics, Uttaranchal College of Science and Technology, Dehradun, INDIA Forest Pathology Division, Forest Research Institute, Dehradun, INDIA Available online at: www.isca.in Received 17th March 2013, revised 5th April 2013, accepted 30th May 2013Abstract In the present investigation, total twenty different protein sequences of rhodopsin from different organisms of vertebrates were obtained from GenPept database and only 347 characters of each sequence were considered for motif discovery, motif family analysis and phylogenetic analysis. Three different motifs were discovered by MEME program. The Pfam analysis of these motifs result revealed that two motifs belonged to 7 transmembrane receptor family. Two major clusters of all retrieved sequences were obtained after phylogenetic analysis. Keywords: Evolutinary analysis, Motif discovery, rhodopsin, vertebrates, GenPept. Introduction Rhodopsins are biological pigments in photoreceptor cells of the retina that is responsible for the first events in the perception of light. Rhodopsin belongs to G-protein-coupled receptor family and is extremely sensitive to light, enabling vision in low-light conditions. Exposed to light, the pigment immediately photobleaches, and it takes about 45 minutes to regenerate fully in humans. Rhodopsin consists of the protein moiety opsin and a reversibly covalently bound cofactor, retinal. Opsin, a bundle of seven transmembrane helices connected to each other by protein loops, binds retinal, which is located in a central pocket on the seventh helix at a lysine residue. Retina lies horizontally with relation to the membrane. Each outer segment disc contains thousands of visual pigment molecules. Retinol is produced in the retina from Vitamin A, from dietary beta-carotene. Isomerization of 11-cis-retinal into all-trans-retinal by light induces a conformational change (bleaching) in opsin, continuing with metarhodopsin II, which activates the associated G protein transducin and triggers a Cyclic Guanosine Monophosphate, second messenger and cascade3,4. Considering the above facts the study of amino acid sequences of rhodopsin from different organisms of vertebrates is quit challenging. In this communication we performed the In silico analysis including motif identification, pfam analysis and phylogenetic analysis of various sequences of rhodopsin from vertebrates. Material and MethodsTwenty different protein sequences of rhodopsin from different organisms of vertebrates were searched in GenPept database and randomly selected. All the selected sequences were opened in MEGA4 program and trimmed from end positions to make them for equal length. Total 347 characters were taken for phylogenetic analysis and motif discovery. Motifs were identified in sequences using MEME program. All motifs were subjected to their family analysis using Pfam Database. The Multiple Sequence Alignment was performed using CLASTAL-W program before phylogenetic tree construction. The phylogenetic analysis was performed by UPGMA method using MEGA4 program. Results and DiscussionAll retrieved sequences along with their accession number, species name and origin is listed in table-1. All the sequences were trimmed from end side to make them for equal lengths. Motif discovery result revealed that three motifs were discovered (figure-1). Figure-2, figure-4 and figure-6 are showing the sites of bock one, two and three respectively. Figure-3, figure-5 and figure-7 are showing the locations of motif one, two and three in each rhodopsin sequences. Figure-8 is showing the combined block diagram all Motifs locations of each block. The Pfam analysis showed that motif second and third was belonging to 7 transmembrane receptor family. The Pfam entry of first motif was not found. The multiple sequence alignment result showed some conserved regions in all aligned sequences. Two major sequences clusters were obtained by phylogenetic analysis. Cluster I consisted of 15 species and further divided in two subcluters. Subcluster I consisted of 11 species namely Rattus norvegicus, Mus musculus, Canis lupus familiaris, Felis catus, Cricetulus griseus, Homo sapiens, Sus scrofa, Cavia porcellus, Bos taurus, Gallus gallus and Taeniopygia guttata. Subcluster II consisted of 4 species namely Xenopus laevis, Xenopus tropicalis, Rana pipiens and Rana catesbeiana. Cluster II contained 5 species namely Sargocentron tiere, Plecoglossus altivelis, Poecilia reticulata, Danio rerio and Cyprinus carpio. International Research Journal of Biological Sciences ________________________________________________ ISSN 2278-3202 Vol. 2(7), 6-11, July (2013) Int. Res. J. Biological Sci. International Science Congress Association 7 Table-1 Organism name and accession number of all retrieved sequences from GenPept of rhodopsin from vertebrates S.No. Organism name Accession number 1.Plecoglossus altivelis BAB88652.1 2.Homo sapiens NP_000530.1 3.Bos taurus NP_001014890.1 4.Rattus norvegicus NP_254276.1 5.Xenopus laevis NP_001080517.1 6.Sus scrofa NP_999386.1 7.Canis lupus familiaris NP_001008277.1 8.Xenopus tropicalis NP_001090803.1 9.Danio rerio NP_571159.1 10.Gallus gallus NP_001025777.1 11.Taeniopygia guttata NP_001070163.1 12.Rana pipiens AAB24265.1 13.Mus musculus NP_663358.1 14.Cavia porcellus NP_001166556.1 15.Cyprinus carpio AAB33306.1 16.Rana catesbeiana AAB35478.1 17.Poecilia reticulata ABI33107.1 18.Felis catus NP_001009242.1 19.Cricetulus griseus NP_001231336.1 20.Sargocentron tiere AAB39534.1 Figure-1 Conserved motifs of rhodopsin International Research Journal of Biological Sciences ________________________________________________ ISSN 2278-3202 Vol. 2(7), 6-11, July (2013) Int. Res. J. Biological Sci. International Science Congress Association 8 Figure-2 Site of Block one Figure-3 Block One Show the Motif Location in each rhodopsin sequencesFigure-4 Site of Block Two International Research Journal of Biological Sciences ________________________________________________ ISSN 2278-3202 Vol. 2(7), 6-11, July (2013) Int. Res. J. Biological Sci. International Science Congress Association 9 Figure-5 Block Two Show the Motif Location in each rhodopsin sequencesFigure-6 Site of Block ThreeFigure-7 Block Three Show the Motif Location in each rhodopsin sequences International Research Journal of Biological Sciences ________________________________________________ ISSN 2278-3202 Vol. 2(7), 6-11, July (2013) Int. Res. J. Biological Sci. International Science Congress Association 10 Figure- 8 Combined block diagram show the Motif location of each blockFigure-9 Phylogenetic analysis of all retrieved sequences of rhodopsin from vertebrates Rattus_norvegicus Mus_musculus Canis_lupus_familiaris Felis_catus Cricetulus_griseus Homo_sapiens Sus_scrofa Cavia_porcellus Bos_taurus Gallus_gallus Taeniopygia_guttata Xenopus_laevis Xenopus_tropicalis Rana_pipiens Rana_catesbeiana Sargocentron_tiere Plecoglossus_altivelis Poecilia_reticulata Danio_rerio Cyprinus_carpio 0.00 0.02 0.04 0.06 0.08 0.10 0.12 International Research Journal of Biological Sciences ________________________________________________ ISSN 2278-3202 Vol. 2(7), 6-11, July (2013) Int. Res. J. Biological Sci. International Science Congress Association 11 Conclusion Motifs identification in a group of related sequences of rhodopsin showed the evolutionary relationships of functional features among different organisms of vertebrates. Three motifs belonging to 7 transmembrane receptor family was identified. This suggests that these motifs have an important function in the evolution of rhodopsin in vertebrates. Two major sequence clusters were obtained by phylogenetic analysis. This suggests that the sequences of cluster I is more closely related in comparison to sequences of cluster II. This classification can significantly contribute in the understanding of the evolutionary relationships between the species at sequence level9-11. References1.Humphries P., Kenna P. and Farrar G.J., On the molecular genetics of retinitis pigmentosa, Science, 256 (5058), 804–(1992) 2.Edwards S.C., Involvement of cGMP and calcium in the photoresponse in vertebrate photoreceptor cells, The Journal of the Florida Medical Association, 82(7), 485–8 (1995)3.Maghtheh M., Gregory C., Inglehearn C., et al., Rhodopsin mutations in autosomal dominant retinitis pigmentosa, Hum. Mutat.,2(4), 249–55 (1993) 4.Garriga P., Manyosa J., The eye photoreceptor protein rhodopsin. Structural implications for retinal disease, FEBS Lett., 528(1–3), 17–22 (2002) 5.Kumar S., Dudley J., Nei M., and Tamura K.,MEGA: a biologist-centric software for evolutionary analysis ofDNAand protein sequences, Briefings in Bioinformatics, , 299-306 (2008) 6.Bailey T.L. and Elkan C., Unsupervised learning of multiple motifs in biopolymers using expectation maximization, Mach Learn, 21(51), 80-33 (1995) 7. Punta M., Coggill P.C., Eberhardt R.Y., Mistry J., Tate J., Boursnell C., Pang N., Forslund K., Ceric G., Clements J., Heger A., Holm L., Sonnhammer E.L.L., Eddy S.R., Bateman A., and Finn R.D. The Pfam Protein Families Database, Nucleic Acids Research Database (2012) 8. Thompson J.D., Higgins D.G., Gibson T.J., CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position specific gap penalties and weight matrix choice, Nucleic Acids Res., 22(22), 4673-80 (1994) 9. Dwivedi V.D., Arora S., Kumar A. and Mishra S.K., Computational analysis of xanthine dehydrogenase enzyme from different source organisms, Network Modeling Analysis in Health Informatics and Bioinformatics, DOI : 10.1007/s13721-013-0029-7(2012) 10. Dhar D.V., Tanuj S., Kumar M.S. and Kumar P.A., Insights to Sequence Information of Lactoylglutathione Lyase Enzyme from Different Source Organisms, I. Res. J. Biological Sci., 1(6), 38-42 (2012) 11.Dhar D.V., Tanuj S., Amit P. and Kumar M.S., Insights to Sequence Information of Alpha Amylase Enzyme from Different Source Organisms, International Journal of Advanced Biotechnology and Bioinformatics, 1(1), 87-91 (2012)