Note to users. If you're seeing this message, it means that your browser cannot find this page's style/presentation instructions -- or possibly that you are using a browser that does not support current Web standards. Find out more about why this message is appearing, and what you can do to make your experience of our site the best it can be.
Subscribe

Sci. STKE, 27 June 2000
Vol. 2000, Issue 38, p. pl1
[DOI: 10.1126/scisignal.382000pl1]

PROTOCOLS

Measurement of Molecular Interactions in Living Cells by Fluorescence Resonance Energy Transfer Between Variants of the Green Fluorescent Protein

Richard M. Siegel1, Francis Ka-Ming Chan1, David A. Zacharias3, Ruth Swofford2, Kevin L. Holmes2, Roger Y. Tsien3, and Michael J. Lenardo1*

1The Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
2The Flow Cytometry Section, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
3The Howard Hughes Medical Institute and Department of Pharmacology, UCSD School of Medicine, La Jolla, CA 92093-0647, USA.

Abstract: Many signal transduction pathways operate through oligomerization of proteins into multi-subunit complexes. Although biochemical assays can identify potential protein-protein interactions, studying these interactions in living cells is more challenging. Fluorescence resonance energy transfer (FRET) has been used as a "spectroscopic ruler" to measure molecular proximity, but these methods have been limited by the need for chemical labeling of target proteins or labeled antibodies. We present methods for examining interactions between target proteins molecularly fused to cyan and yellow variants of the green fluorescent protein (GFP) by FRET in living cells. Flow cytometric and microscope-based methods are described that have been applied to a variety of interacting proteins.

*Corresponding author, E-mail: lenardo{at}nih.gov

Citation: R. M. Siegel, F. K.-M. Chan, D. A. Zacharias, R. Swofford, K. L. Holmes, R. Y. Tsien, M. J. Lenardo, Measurement of Molecular Interactions in Living Cells by Fluorescence Resonance Energy Transfer Between Variants of the Green Fluorescent Protein. Sci. STKE 2000, pl1 (2000).

Read the Full Text

THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES:
Orai1 Determines Calcium Selectivity of an Endogenous TRPC Heterotetramer Channel.
D. L. Cioffi, S. Wu, H. Chen, M. Alexeyev, C. M. St. Croix, B. R. Pitt, S. Uhlig, and T. Stevens (2012)
Circ. Res. 110, 1435-1444
   Abstract »    Full Text »    PDF »
The PSAP Motif within the ORF3 Protein of an Avian Strain of the Hepatitis E Virus Is Not Critical for Viral Infectivity In Vivo but Plays a Role in Virus Release.
S. P. Kenney, R. S. Pudupakam, Y.-W. Huang, F. W. Pierson, T. LeRoith, and X.-J. Meng (2012)
J. Virol. 86, 5637-5646
   Abstract »    Full Text »    PDF »
Regulation of phenotypic variability by a threshold-based mechanism underlies bacterial persistence.
E. Rotem, A. Loinger, I. Ronin, I. Levin-Reisman, C. Gabay, N. Shoresh, O. Biham, and N. Q. Balaban (2010)
PNAS 107, 12541-12546
   Abstract »    Full Text »    PDF »

To Advertise     Find Products

Science Signaling. ISSN 1937-9145 (online), 1945-0877 (print). Pre-2008: Science's STKE. ISSN 1525-8882