A novel enzyme complementation-based assay for monitoring G-protein-coupled receptor internalization.

A novel enzyme complementation-based assay for monitoring G-protein-coupled receptor internalization. Research Abstract Details 

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A novel enzyme complementation-based assay for monitoring G-protein-coupled receptor internalization. Abstract Text:

Mark M Hammer,Tom S Wehrman,Helen M Blau,Mark M Hammer,Tom S Wehrman,Helen M Blau,

G-protein-coupled receptor (GPCR) signaling is involved in a wide range of physiological processes and diseases, and around one-half of currently used drugs target GPCRs. Assays for the signaling of GPCRs have suffered from drawbacks, including low signal-to-noise, temporally transient signals, and difficulty in applying a single assay to a wide range of GPCRs. We have developed a set of assays for G-protein-coupled receptor signaling based on beta-galactosidase enzyme complementation in live mammalian cells. We previously described an assay for GPCR activation by monitoring the binding of beta-arrestin to the receptor. Here we describe a second assay that monitors the internalization of GPCRs to endosomes, an event that follows receptor activation and is critical in desensitizing and resensitizing the receptor. We show that both assays display high signal-to-noise ratios with low variability and are quantitative for a wide range of GPCRs. EC50s obtained with these assays closely match results reported in the literature. Finally, we show that these assays are readily adapted to high-throughput chemical screens. Thus, these two assays for monitoring G-protein-coupled receptor activation and internalization should prove valuable in basic biological studies as well as in high-throughput screens.

A novel enzyme complementation-based assay for monitoring G-protein-coupled receptor internalization. Publishing Authors By Initials

MM Hammer,TS Wehrman,HM Blau,MM Hammer,TS Wehrman,HM Blau,

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A novel enzyme complementation-based assay for monitoring G-protein-coupled receptor internalization. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: The FASEB journal : official publication of the Fe

VOLUME: 21

Page Numbers: 3827-34

Journal Abbreviation: FASEB J.

ISSN: 1530-6860

DAY: 17

MONTH: 10

YEAR: 2007

A novel enzyme complementation-based assay for monitoring G-protein-coupled receptor internalization. Information

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LANGUAGE: eng

NlmUniqueID: 8804484

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Grant and Affiliation Information for A novel enzyme complementation-based assay for monitoring G-protein-coupled receptor internalization.

AFFILIATION: Baxter Laboratory in Genetic Pharmacology, Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California 94305-5175, USA.

Country: United States

AGENCY: United States NIGMS

GRANT: T32 GM08412

ACRONYM: GM

MEDLINETA: FASEB J

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