title: Analysis of Conserved Glutamate and Aspartate Residues in Drosophila Rhodopsin 1 and Their Influence on Spectral Tuning.
abstract: The molecular mechanisms that regulate invertebrate visual pigment absorption are poorly understood. Studies of amphioxus Go-opsin have demonstrated that Glu-181 functions as the counterion in this pigment. This finding has led to the proposal that Glu-181 may function as the counterion in other invertebrate visual pigments as well. Here we describe a series of mutagenesis experiments to test this hypothesis and to also test whether other conserved acidic amino acids in Drosophila Rhodopsin 1 (Rh1) may serve as the counterion of this visual pigment. Of the 5 Glu and Asp residues replaced by Gln or Asn in our experiments, none of the mutant pigments shift the absorption of Rh1 by more than 6 nm. In combination with prior studies, these results suggest that the counterion in Drosophila Rh1 may not be located at Glu-181 as in amphioxus, or at Glu-113 as in bovine rhodopsin. Conversely, the extremely low steady state levels of the E194Q mutant pigment (bovine opsin site Glu-181), and the rhabdomere degeneration observed in flies expressing this mutant demonstrate that a negatively charged residue at this position is essential for normal rhodopsin function in vivo. This work also raises the possibility that another residue or physiologic anion may compensate for the missing counterion in the E194Q mutant.
authors.affiliation: From the Departments of Cell and Developmental Biology.
authors.forename: Lijun
authors.initials: L
authors.lastname: Zheng
authors.affiliation: Pharmacology, and.
authors.forename: David M
authors.initials: DM
authors.lastname: Farrell
authors.affiliation: From the Departments of Cell and Developmental Biology.
authors.forename: Ruth M
authors.initials: RM
authors.lastname: Fulton
authors.affiliation: From the Departments of Cell and Developmental Biology.
authors.forename: Eve E
authors.initials: EE
authors.lastname: Bagg
authors.affiliation: From the Departments of Cell and Developmental Biology.
authors.forename: Ernesto
authors.initials: E
authors.lastname: Salcedo
authors.affiliation: From the Departments of Cell and Developmental Biology.
authors.forename: Meridee
authors.initials: M
authors.lastname: Manino
authors.affiliation: From the Departments of Cell and Developmental Biology, Ophthalmology and Rocky Mountain Lions Eye Institute, University of Colorado, Anschutz Medical Campus, School of Medicine, Aurora, Colorado 80045 steve.britt@ucdenver.edu.
authors.forename: Steven G
authors.initials: SG
authors.lastname: Britt
mesh_terms: D000818:Animals; D001224:Aspartic Acid / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015153:Blotting, Western; D017124:Conserved Sequence / Q000235:genetics; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism; D018698:Glutamic Acid / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015591:Microspectrophotometry; D009154:Mutation; D055355:Opsins / Q000145:classification / Q000235:genetics / Q000378:metabolism; D010802:Phylogeny; D017433:Protein Structure, Secondary; D012162:Retinal Degeneration / Q000235:genetics / Q000378:metabolism; D012243:Rhodopsin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D029721:Drosophila Proteins; D055355:Opsins; D001224:Aspartic Acid; D018698:Glutamic Acid; D012243:Rhodopsin
keywords: Drosophila; color perception; photoreceptor; retina; rhodopsin; site-directed mutagenesis