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Functional characterization of rhodopsin monomers and dimers in detergents

Jastrzebska, B. and Maeda, T. and Zhu, L. and Fotiadis, D. and Filipek, S. and Engel, A. and Stenkamp, R. E. and Palczewski, K.. (2004) Functional characterization of rhodopsin monomers and dimers in detergents. Journal of biological chemistry, Vol. 279. pp. 54663-54675.

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Official URL: http://edoc.unibas.ch/dok/A5262409

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Abstract

Rhodopsin (Rho) is a G protein-coupled receptor that initiates phototransduction in rod photoreceptors. High expression levels of Rho in the disc membranes of rod outer segments and the propensity of Rho to form higher oligomeric structures are evident from atomic force microscopy, transmission electron microscopy, and chemical crosslinking experiments. To explore the structure and functional properties of Rho in n-dodecyl-ss-maltoside (DM), frequently used to purify heterologously expressed Rho and its mutants, we used gel filtration techniques, blue native gel electrophoresis, and functional assays. Here, we show that in micelles containing DM at concentrations greater than 3 mM, Rho is present as a single monomer per detergent micelle. In contrast, in 12 mM 3-[(3-cholamidopropyl)dimethyl-ammonio]-1-propanesulfonate (CHAPS), micelles contain mostly dimeric Rho. The cognate G protein transducin (Gt) appears to have a preference for binding to the Rho dimer, and the complexes fall apart in the presence of GTPS. Crosslinked Rho dimers release the chromophore at a slower rate than monomers and are much more resistant to heat denaturation. Both Rho* monomers and dimers are capable of activating Gt and both of them are phosphorylated by Rho kinase. Rho expressed in HEK293 cells is also readily crosslinked by a bi-functional reagent. These studies provide an explanation of how detergent influences the oligomer-dimer-monomer equilibrium of Rho and describe the functional characterization of Rho monomers and dimers in detergent.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Structural Biology (Engel)
UniBasel Contributors:Engel, Andreas H
Item Type:Article, refereed
Article Subtype:Research Article
Bibsysno:Link to catalogue
Publisher:American Society of Biological Chemists
ISSN:0021-9258
Note:Publication type according to Uni Basel Research Database: Journal article
Last Modified:22 Mar 2012 14:21
Deposited On:22 Mar 2012 13:23

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