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PMID: 2962193 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Expression of a synthetic bovine rhodopsin gene in monkey kidney cells.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 84 ·No. 24 ·1987-12-00 ·Pages 8874-8

Oprian DD, Molday RS, Kaufman RJ, Khorana HG

Abstract

We report here the high-level expression of a synthetic gene for bovine rhodopsin in transfected monkey kidney COS-1 cells. Rhodopsin is produced in these cells to a level of 0.3% of the cell protein, and it binds exogenously added 11-cis-retinal to generate the characteristic rhodopsin absorption spectrum. We describe a one-step immunoaffinity procedure for purification of the rhodopsin essentially to homogeneity. The COS-1 cell rhodopsin activates the GTPase activity of bovine transducin in a light-dependent manner with the same specific activity as that of purified bovine rhodopsin. Electron microscopy of immunogold-stained cells indicates that rhodopsin is located in the plasma membrane of the transfected cells and is oriented with the amino terminus on the extracellular side of the membrane. This orientation is analogous to that of rhodopsin in the disk membranes of photoreceptor cells in the bovine retina.

MeSH Terms
Animals Cattle Cell Compartmentation Cell Membrane/metabolism Chlorocebus aethiops DNA, Recombinant Eye Proteins/genetics Immunohistochemistry Light Retinal Pigments/genetics Retinaldehyde/metabolism Rhodopsin/genetics Rod Opsins Spectrum Analysis Transfection
Chemicals
DNA, Recombinant Eye Proteins Retinal Pigments Rod Opsins Rhodopsin Retinaldehyde
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Oprian D D
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Molday R S
Kaufman R J
Khorana H G
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.84.24.8874
Published
1987-12-00
Pages
8874-8
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC299653
Subset
IM
Grants
NEI NIH HHS · EY-02422 · United States
NIGMS NIH HHS · GM28289-07 · United States
External Links
PubMed source DOI: 10.1073/pnas.84.24.8874 PMC Full Text View journal details
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