Cones must function over a wide range of light intensities and a broad span of the electromagnetic spectrum. The ability to perform under such diverse conditions arises in large part from the unique set of phototransduction proteins found in cone cells. Light responses initiate with the absorption of photons by the visual pigments (cone opsins). The long term goals of this project are to understand how the cone opsin family has attained absorption maxima from 350 nm to 650 nm while achieving the rapid photobleaching and regeneration properties that play a large role in shaping the light responses of cones. This proposal will combine molecular, biochemical and biophysical approaches to investigate the molecular mechanisms underlying the function of cone visual pigments. In our First Aim, the short wavelength-sensitive (SWS1) visual pigments will be purified from transfected COS cells in order to investigate how particular amino acid residues in the transmembrane helical segments and the extracellular domain regulate photoactivation and regeneration properties. The conformational changes that occur following isomerization of 11-cis retinal will be studied by low temperature UV/visible and fluorescence spectroscopy. The amino acids that regulate the formation and stability of the light-activated conformation (R*) will be studied in conjunction with molecular modeling. These experiments will allow us to address in a comprehensive way several long standing issues about visual pigment behavior: mechanisms controlling light-activation and pigment regeneration. In order to study the structure of cone visual pigments, in our Second Aim we will investigate in a systematic fashion the stability of cone opsins in a variety of solubilizing agents. To accomplish this we will use an inducible heterologous over-expression system to produce milligram quantities of cone pigment. The information gained in this Aim will allow for protein crystallization trials in the future.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY011256-11
Application #
7998169
Study Section
Biology and Diseases of the Posterior Eye Study Section (BDPE)
Program Officer
Neuhold, Lisa
Project Start
1996-12-01
Project End
2012-11-30
Budget Start
2010-12-01
Budget End
2012-11-30
Support Year
11
Fiscal Year
2011
Total Cost
$298,426
Indirect Cost
Name
Upstate Medical University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
058889106
City
Syracuse
State
NY
Country
United States
Zip Code
13210
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Adekeye, Adeseye; Haeri, Mohammad; Solessio, Eduardo et al. (2014) Ablation of the proapoptotic genes CHOP or Ask1 does not prevent or delay loss of visual function in a P23H transgenic mouse model of retinitis pigmentosa. PLoS One 9:e83871
Sandberg, Megan N; Greco, Jordan A; Wagner, Nicole L et al. (2014) Low-Temperature Trapping of Photointermediates of the Rhodopsin E181Q Mutant. SOJ Biochem 1:
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Haeri, Mohammad; Calvert, Peter D; Solessio, Eduardo et al. (2013) Regulation of rhodopsin-eGFP distribution in transgenic xenopus rod outer segments by light. PLoS One 8:e80059
Zhuo, Xinming; Haeri, Mohammad; Solessio, Eduardo et al. (2013) An inducible expression system to measure rhodopsin transport in transgenic Xenopus rod outer segments. PLoS One 8:e82629
Kuemmel, Colleen M; Sandberg, Megan N; Birge, Robert R et al. (2013) A conserved aromatic residue regulating photosensitivity in short-wavelength sensitive cone visual pigments. Biochemistry 52:5084-91
Zuber, Michael E; Nihart, Heather S; Zhuo, Xinming et al. (2012) Site-specific transgenesis in Xenopus. Genesis 50:325-32
Haeri, Mohammad; Knox, Barry E (2012) Rhodopsin mutant P23H destabilizes rod photoreceptor disk membranes. PLoS One 7:e30101
Haeri, Mohammad; Knox, Barry E (2012) Generation of transgenic Xenopus using restriction enzyme-mediated integration. Methods Mol Biol 884:17-39

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