This Article Full Text Full Text (PDF) All Versions of this Article: 19/7/1821    most recent Author Manuscript (PDF) Purchase Article View Shopping Cart Alert me when this article is cited 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 Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Dong, M. Articles by Miller, L. J. Search for Related Content PubMed PubMed Citation Articles by Dong, M. Articles by Miller, L. J.

Insights into the Structure and Molecular Basis of Ligand Docking to the G Protein-Coupled Secretin Receptor Using Charge-Modified Amino-Terminal Agonist Probes

Cancer Center and Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic Scottsdale, Scottsdale, Arizona 85259

Address all correspondence and requests for reprints to: Laurence J. Miller, M.D. Director, Cancer Center Mayo Clinic in Scottsdale, 13400 East Shea Boulevard, Johnson Research Building, Scottsdale Arizona 85259. E-mail: miller{at}mayo.edu.

The amino terminus and third loop regions of class B G protein-coupled receptors play critical roles in ligand docking and action. For the prototypic secretin receptor, the hormone amino terminus is spatially approximated with receptor region high in transmembrane segment 6 (TM6), whereas residues ranging from position 6 through 26 label the amino terminus. Here, we focus on the role of charge of the secretin amino terminus, using a series of full-agonist, acetylated probes. Sites of covalent labeling were examined using sequential purification, chemical and enzymatic cleavage, and Edman degradation. High-affinity amino-terminally-blocked probes labeled the distal amino-terminal tail, rather than TM6, while adding a basic residue, again labeled TM6. These data suggest that the secretin amino terminus docks between the amino terminus and TM6 of the receptor, with this region of secretin likely interacting with an acidic residue within the receptor TM6 and the third extracellular loop. To explore this, candidate acidic residues were mutated to Ala (E341A, D342A, E345A, E351A). The E351A mutant markedly interfered with binding, biological activity, and internalization, whereas all others bound secretin and signaled and internalized normally. This supports the possibility that there is a charge-charge interaction between this residue and the amino terminus of secretin that is critical to its normal docking.

This article has been cited by other articles:

				S. Runge, H. Thogersen, K. Madsen, J. Lau, and R. Rudolph Crystal Structure of the Ligand-bound Glucagon-like Peptide-1 Receptor Extracellular Domain J. Biol. Chem., April 25, 2008; 283(17): 11340 - 11347. [Abstract] [Full Text] [PDF]

				K. G. Harikumar, D. I. Pinon, and L. J. Miller Transmembrane Segment IV Contributes a Functionally Important Interface for Oligomerization of the Class II G Protein-coupled Secretin Receptor J. Biol. Chem., October 19, 2007; 282(42): 30363 - 30372. [Abstract] [Full Text] [PDF]

				C. S. Lisenbee, K. G. Harikumar, and L. J. Miller Mapping the Architecture of Secretin Receptors with Intramolecular Fluorescence Resonance Energy Transfer Using Acousto-Optic Tunable Filter-Based Spectral Imaging Mol. Endocrinol., August 1, 2007; 21(8): 1997 - 2008. [Abstract] [Full Text] [PDF]

				T. Dean, A. Khatri, Z. Potetinova, G. E. Willick, and T. J. Gardella Role of Amino Acid Side Chains in Region 17-31 of Parathyroid Hormone (PTH) in Binding to the PTH Receptor J. Biol. Chem., October 27, 2006; 281(43): 32485 - 32495. [Abstract] [Full Text] [PDF]

				M. Beinborn Class B GPCRs: A Hidden Agonist Within? Mol. Pharmacol., July 1, 2006; 70(1): 1 - 4. [Abstract] [Full Text] [PDF]

				M. Dong, D. I. Pinon, Y. W. Asmann, and L. J. Miller Possible Endogenous Agonist Mechanism for the Activation of Secretin Family G Protein-Coupled Receptors Mol. Pharmacol., July 1, 2006; 70(1): 206 - 213. [Abstract] [Full Text] [PDF]