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Use of a distant reporter group as evidence for a conformational change in a sensory receptor

Zukin, R. S. and Hartig, P. R. and Koshland, D. E., Jr. (1977) Use of a distant reporter group as evidence for a conformational change in a sensory receptor. Proceedings of the National Academy of Sciences of the United States of America, 74 (5). pp. 1932-1936. ISSN 0027-8424.

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A highly sensitive method for demonstrating ligand-induced conformational changes in protein molecules in solution is described. The method utilizes an environmentally sensitive reporter group that is known to be distant from the active site. In the present application a conformational change is demonstrated in the galactose receptor of Salmonella typhimurium, involved in bacterial sensing and transport, by means of an extrinsic fluorophore, 5-iodoacetamidofluorescein, attached at a single methionine residue, and the intrinsic tryptophan fluorophore. Binding of the ligand galactose perturbs the microenvironment of both the fluorescein and tryptophan, as shown by both spectral and potassium iodide quenching changes. The distance between the two dyes is established by fluorescence energy transfer methods to be 41 ± 10 angstrom. Since only one molecule of galactose binds per molecule of receptor and since the galactose molecule is only about 5 angstrom in length, changes at one of these sites reflect the result of an indirect effect. Hence, there must be a ligand-induced conformational change that is propagated a minimum of 30 angstrom through the receptor molecule.

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Additional Information:© 1977 by the National Academy of Sciences. Contributed by Daniel E. Koshland, Jr., February 28, 1977. We thank Dr. Terry Leighton for assistance and advice with peptide chromatography and Ms. Judy Benson and Dr. M.E. Koshland for assistance and advice in the amino-acid analyses. The work was funded by U.S. Public Health Service Grant AM9765 and National Science Foundation Grant PCM76-21850. The costs of publication of this article were defrayed in part by the payment of page charges from funds made available to support the research which is the subject of the article. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Subject Keywords:galactose receptor; 5-iodoacetamidofluorescein; chemotaxis; spectroscopic rules
Issue or Number:5
Record Number:CaltechAUTHORS:ZUKpnas77
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Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:9481
Deposited By: Tony Diaz
Deposited On:23 Jan 2008
Last Modified:03 Oct 2019 00:00

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