. Role of key aromatic residues in the ligand-binding domain of alpha7 nicotinic receptors in the agonist action of beta-amyloid. J Biol Chem. 2011 Sep 30;286(39):34373-81. PubMed.


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  1. This study by Tong et al. provides the first description of amino acid residues that contribute (or not) to the activation of the homomeric α7 nicotinic acetylcholine receptor (nAChR) by β amyloid (Aβ). Given the mounting evidence that α7 nAChRs and oligomeric Aβ undergo prolonged interaction, and possibly provide a scaffold for Aβ accumulation in Alzheimer’s disease (AD), these findings have exciting implications for therapeutic interventions that would target the interaction of these two proteins.

    Employing the rodent hybrid neuroblastoma cell line, NG108-15, as a heterologous expression system, and calcium imaging to detect receptor activation, the Nichols’ group first established that the extracellular domain of the α7 nAChR is necessary and sufficient for Aβ-induced intracellular calcium increases. The researchers expressed a chimeric receptor in which the transmembrane and intracellular domains of the α7 nAChR were replaced with a sequence from the homomeric serotonergic ligand-gated ion channel that is unresponsive to Aβ.

    Next, they mutated key aromatic residues in the three binding loops (A, B, and C) previously identified as important for agonist activation of this class of receptors. The mutation that most affected responsiveness to Aβ was tyrosine 188, situated in the “aromatic cluster” of loop C of the extracellular agonist binding domain. The other aromatic residues mutated in the ligand-binding domain were tyrosine 195 (loop C), tyrosine 93 (loop A), and tryptophan 149 (loop B). These other mutations had minimal effect on acetylcholine (ACh) or Aβ receptor activation. Since these residues have previously been implicated in the binding and activation of the receptor by nicotine, but not ACh, it suggests that Aβ interacts with the α7 nAChR in an analogous manner to ACh (O'Leary and White, 1992).

    To further test this idea, Tong et al. mutated tyrosine 188 both to a serine and a phenylalanine. The tyrosine to serine mutation results in loss of the tyrosine aromatic group, whereas tyrosine to phenylalanine results in loss of the hydroxyl group. The observation that either mutation resulted in significant attenuation of Aβ- and ACh-induced calcium responses suggests that Aβ binding, similar to that of ACh, is stabilized by both the hydroxyl and aromatic groups of tyrosine 188. Previous work has established this residue as differentially interacting with the quaternary amine of ACh in contrast to nicotine (Celie et al., 2004). As one might predict, the Nichols’ group found that mutation of tyrosine 188 to either phenylalanine or serine led to near complete loss of receptor responses to Aβ and ACh, but not to nicotine, demonstrating that this residue is essential for the agonist-like action of Aβ.

    Finally, the group employed immunostaining and biochemical analyses to detect surface receptors and demonstrated that receptor-peptide complexes were present on the presynaptic-like varicosities formed by this cell line in culture, thus providing additional evidence that functional changes resulting from receptor mutagenesis were likely due to Aβ-α7 nAChR interactions at the presynaptic cell surface.

    There are several additional aspects to this study that are noteworthy. First, the effective concentration of Aβ on the α7 nAChRs studied ranged from picomolar to nanomolar, considered to be within the physiological range for the amyloid peptide (Kuo et al., 1996). Second, the Aβ preparation employed has been previously characterized as comprising mainly tetrameric and hexameric peptide assemblies, based on molecular weight standards under non-denaturing gel electrophoresis conditions (Khan et al., 2010). Third, the results further suggest a potential cation-π interaction between Aβ and tyrosine 188 in the α7-nAChR, given the similarity in the effect of the mutations on receptor activation by ACh and Aβ. This further suggests that basic amino acid residue(s) and/or aromatic residue(s) within the hydrophilic region of Aβ (~1-28) contribute to this interaction. Fourth, it is possible that a portion of the Aβ peptide may actually insert itself into the agonist binding pocket, since α-bungarotoxin, which is a highly selective 8 kDa peptide antagonist of α7-nAChRs, was found to insert a sequence of amino acid residues (finger II) into this region of the α1 subunit (Dellisanti et al., 2007), coming in close proximity to Y190, which is equivalent to Y188 in the α7 subunit. Last, the Nichols’ group reports that a manuscript in preparation describes studies in which they interrogate the amino acid residues within the Aβ peptide that are necessary and sufficient for agonist activity, and tantalize us with this tidbit: The hydrophilic domain of Aβ (~1-28) is sufficient for its agonist activity.

    Future structural studies will hopefully attempt to refine our understanding of both the structure and stoichiometry of the Aβ oligomers that act on α7 nAChRs as well as the contact points within the peptide-receptor complex. Likewise, it will also be important to pull out the α7 nAChR pharmacological toolkit to investigate if any of the several α7-selective agents affect the binding of Aβ to the receptor and subsequent receptor activation.


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    . Role of key aromatic residues in the ligand-binding domain of alpha7 nicotinic receptors in the agonist action of beta-amyloid. J Biol Chem. 2011 Sep 30;286(39):34373-81. PubMed.

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