Activation of muscarinic acetylcholine receptors via their allosteric binding sites.

Abstract

Ligands that bind to the allosteric-binding sites on muscarinic acetylcholine receptors alter the conformation of the classical-binding sites of these receptors and either diminish or increase their affinity for muscarinic agonists and classical antagonists. It is not known whether the resulting conformational change also affects the interaction between the receptors and the G proteins. We classical-binding site (12-18). The attachment of the allosteric modulator to the allosteric-binding site brings about a change in the conformation of the classical (orthosteric)-binding site and thereby an increase or (more usually) a decrease in the affinity of the receptor for muscarinic agonists and competitive antagonists.The effects of the activation of the allosteric-binding site on the function of the classical-binding site (as reflected by changes in the association, dissociation, and equilibrium binding of muscarinic agonists and competitive antagonists) have been amply investigated (6-8). However, little attention has been paid to the possibility that the allosteric modulators might also affect the conformation and function of the G protein binding site and, consequently, the interaction between muscarinic receptors, G proteins, and the G protein-regulated effector molecules. We present evidence that the allosteric modulators alcuronium, gallamine, and strychnine acting in the absence of agonists have profound agonist-like effects on the synthesis of cAMP and inositol phosphates (IPs) in cells stably transfected with the genes for muscarinic receptors and that, apparently, muscarinic receptors can be activated not only by ligands acting via their classical-binding site, but also by ligands acting via the allosteric-binding site. We suggest that the same novel mechanism of receptor activation may operate on the other G protein-coupled receptors.Muscarinic acetylcholine receptors belong to the large group of receptors coupled with G proteins (1). Signal transduction from the M2 and M4 muscarinic receptor subtypes occurs preferentially via the guanine nucleotide-binding regulatory proteins Gi and Go, whereas the M1, M3, and M5 receptor subtypes couple preferentially with proteins of the Gq family (2, 3). Muscarinic agonists bind to the classical-binding site, which is located in the depth of the cell membrane in a barrel formed by the seven transmembrane segments of the receptor molecule (4, 5). Agonist binding brings about a change in the conformation of the intracellular receptor domain responsible for the contact with the G protein, and this change brings about the activation of the G protein. Although details of these interactions are the subject of scrutiny and debate, it is generally believed that the attachment of the signal molecule to the classical-binding site is the necessary first step in muscarinic receptor-mediated signal transduction across the cell membrane.However, it is well known that the binding properties of the classical-binding sites of muscarinic receptors are subject to allosteric reg...

Wash-Resistantly Bound Xanomeline Inhibits Acetylcholine Release by Persistent Activation of Presynaptic M2and M4Muscarinic Receptors in Rat Brain

“…Interaction of xanomeline at an allosteric site on the muscarinic receptor is probably involved in its wash-resistant binding and agonistic effects (Jakubík et al, 2002). Thus, our findings support the potential of allosteric agonists (Lazareno and Birdsall, 1995;Jakubík et al, 1996Jakubík et al, , 1998Jakubík et al, , 2006Sur et al, 2003;Langmead et al, 2006), and they provide an example of an agonist drug with prolonged activity that lingers in the absence of free ligand. …”

Small-molecule agonists for the glucagon-like peptide 1 receptor

“…The compounds later were discovered to increase binding of the radiolabeled peptide ligand isotope and, thus, could theoretically have been identified in a binding screen if it had been set up to look for increased tracer binding. The lack of competition with GLP-1 would be consistent with the existence of an allosteric-binding site for the compounds on the GLP-1 receptor, as also described for other small-molecule ligands of GPCRs (33,34), especially the muscarinic and metabotropic glutamate receptors (35,36). However, the compounds described here are also agonists themselves.…”

Positive Cooperativity of Acetylcholine and Other Agonists with Allosteric Ligands on Muscarinic Acetylcholine Receptors

“…Under these conditions, no consistent correlation is apparent between the K L values of agonists in Table 2 and the relevant cooperativity factors ␤ in Table 4. For complete analysis, it will be necessary to explore the cooperativity between agonists and allosteric modulators under conditions in which the receptor/G protein coupling occurs in a normal way, but this will be demanding in view of the complexity of the system and of recent data indicating that the allosteric modulators themselves modify the receptor/G protein interactions (29).…”