HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 282, Issue 35, 25677-25686, August 31, 2007

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 282/35/25677    most recent M702311200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Avlani, V. A. Articles by Christopoulos, A. Search for Related Content PubMed PubMed Citation Articles by Avlani, V. A. Articles by Christopoulos, A. Social Bookmarking                      What's this?

Critical Role for the Second Extracellular Loop in the Binding of Both Orthosteric and Allosteric G Protein-coupled Receptor Ligands^*

Vimesh A. Avlani¹, Karen J. Gregory¹², Craig J. Morton, Michael W. Parker³, Patrick M. Sexton⁴, and Arthur Christopoulos, A Senior Research Fellow of the National Health and Medical Research Council and a Federation Fellow of the Australian Research Council⁵

From the Drug Discovery Biology Laboratory, Department of Pharmacology, Monash University, Clayton, 3800 Australia and Biota Structural Biology Laboratory, St. Vincent's Institute, Fitzroy, 3065, Victoria, Australia

The second extracellular (E2) loop of G protein-coupled receptors (GPCRs) plays an essential but poorly understood role in the binding of non-peptidic small molecules. We have utilized both orthosteric ligands and allosteric modulators of the M₂ muscarinic acetylcholine receptor, a prototypical Family A GPCR, to probe possible E2 loop binding dynamics. We developed a homology model based on the crystal structure of bovine rhodopsin and predicted novel cysteine substitutions that should dramatically reduce E2 loop flexibility via disulfide bond formation and significantly inhibit the binding of both types of ligands. This prediction was validated experimentally using radioligand binding, dissociation kinetics, and cell-based functional assays. The results argue for a flexible "gatekeeper" role of the E2 loop in the binding of both allosteric and orthosteric GPCR ligands.

Received for publication, March 16, 2007 , and in revised form, May 31, 2007.

^* This work was funded by National Health and Medical Research Council of Australia Grant 400134. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Table 1.

¹ These authors contributed equally to the work.

² A recipient of an National Health and Medical Research Council Dora Lush Postgraduate Research Scholarship and a Dowd Foundations Scholarship in the Neurosciences.

³ A Senior Principal Research Fellow of the National Health and Medical Research Council.

⁴ A Principal Research Fellow of the National Health and Medical Research Council.

⁵ To whom correspondence should be addressed. Tel.: 613-9905-3817; Fax: 613-9905-5851; E-mail: arthur.christopoulos{at}med.monash.edu.au.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				C. S. Sum, I. G. Tikhonova, S. Costanzi, and M. C. Gershengorn Two Arginine-Glutamate Ionic Locks Near the Extracellular Surface of FFAR1 Gate Receptor Activation J. Biol. Chem., February 6, 2009; 284(6): 3529 - 3536. [Abstract] [Full Text] [PDF]

				C. Valant, K. J. Gregory, N. E. Hall, P. J. Scammells, M. J. Lew, P. M. Sexton, and A. Christopoulos A Novel Mechanism of G Protein-coupled Receptor Functional Selectivity: MUSCARINIC PARTIAL AGONIST McN-A-343 AS A BITOPIC ORTHOSTERIC/ALLOSTERIC LIGAND J. Biol. Chem., October 24, 2008; 283(43): 29312 - 29321. [Abstract] [Full Text] [PDF]

				L. Zhang, A. Hu, H. Yuan, L. Cui, G. Miao, X. Yang, L. Wang, J. Liu, X. Liu, S. Wang, et al. A Missense Mutation in the CHRM2 Gene Is Associated With Familial Dilated Cardiomyopathy Circ. Res., June 6, 2008; 102(11): 1426 - 1432. [Abstract] [Full Text] [PDF]

				V. A. Avlani, D. J. McLoughlin, P. M. Sexton, and A. Christopoulos The Impact of Orthosteric Radioligand Depletion on the Quantification of Allosteric Modulator Interactions J. Pharmacol. Exp. Ther., June 1, 2008; 325(3): 927 - 934. [Abstract] [Full Text] [PDF]

				F. J. Ehlert and M. T. Griffin Two-State Models and the Analysis of the Allosteric Effect of Gallamine at the M2 Muscarinic Receptor J. Pharmacol. Exp. Ther., June 1, 2008; 325(3): 1039 - 1060. [Abstract] [Full Text] [PDF]

				D. Jager, C. Schmalenbach, S. Prilla, J. Schrobang, A. Kebig, M. Sennwitz, E. Heller, C. Trankle, U. Holzgrabe, H.-D. Holtje, et al. Allosteric Small Molecules Unveil a Role of an Extracellular E2/Transmembrane Helix 7 Junction for G Protein-coupled Receptor Activation J. Biol. Chem., November 30, 2007; 282(48): 34968 - 34976. [Abstract] [Full Text] [PDF]