From ab initio quantum mechanics to molecular neurobiology: a cation-pi binding site in the nicotinic receptor.

The nicotinic acetylcholine receptor is the prototype ligand-gated ion channel. A number of aromatic amino acids have been identified as contributing to the agonist binding site, suggesting that cation-pi interactions may be involved in binding the quaternary ammonium group of the agonist, acetylcholine. Here we show a compelling correlation between: (i) ab initio quantum mechanical predictions of cation-pi binding abilities and (ii) EC50 values for acetylcholine at the receptor for a series of tryptophan derivatives that were incorporated into the receptor by using the in vivo nonsense-suppression method for unnatural amino acid incorporation. Such a correlation is seen at one, and only one, of the aromatic residues-tryptophan-149 of the alpha subunit. This finding indicates that, on binding, the cationic, quaternary ammonium group of acetylcholine makes van der Waals contact with the indole side chain of alpha tryptophan-149, providing the most precise structural information to date on this receptor. Consistent with this model, a tethered quaternary ammonium group emanating from position alpha149 produces a constitutively active receptor.

[1]  D. A. Dougherty,et al.  The Cationminus signpi Interaction. , 1997, Chemical reviews.

[2]  C. Glabe,et al.  Biosynthetic site-specific incorporation of a non-natural amino acid into a polypeptide , 1989 .

[3]  G N Filatov,et al.  The role of conserved leucines in the M2 domain of the acetylcholine receptor in channel gating. , 1995, Molecular pharmacology.

[4]  H. Lester,et al.  Activation of acetylcholine receptor channels by covalently bound agonists in cultured rat myoballs. , 1986, The Journal of physiology.

[5]  Lixin Tang,et al.  Channel gating governed symmetrically by conserved leucine residues in the M2 domain of nicotinic receptors , 1995, Nature.

[6]  G. Tomaselli,et al.  Mutations affecting agonist sensitivity of the nicotinic acetylcholine receptor. , 1991, Biophysical journal.

[7]  D. A. Dougherty,et al.  Cation-π Interactions in Chemistry and Biology: A New View of Benzene, Phe, Tyr, and Trp , 1996, Science.

[8]  A. Karlin,et al.  Toward a structural basis for the function of nicotinic acetylcholine receptors and their cousins , 1995, Neuron.

[9]  D A Dougherty,et al.  Acetylcholine binding by a synthetic receptor: implications for biological recognition , 1990, Science.

[10]  J. Sussman,et al.  Acetylcholinesterase: Structure and use as a model for specific cation-protein interactions , 1992, Current Biology.

[11]  A. Karlin,et al.  Negatively charged amino acid residues in the nicotinic receptor delta subunit that contribute to the binding of acetylcholine. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[12]  A. Goldman,et al.  Atomic structure of acetylcholinesterase from Torpedo californica: a prototypic acetylcholine-binding protein , 1991, Science.

[13]  S. Sharp,et al.  Structure of the agonist-binding site of the nicotinic acetylcholine receptor. [3H]acetylcholine mustard identifies residues in the cation-binding subsite. , 1991, The Journal of biological chemistry.

[14]  D A Dougherty,et al.  Cation-pi interactions in aromatics of biological and medicinal interest: electrostatic potential surfaces as a useful qualitative guide. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[15]  G. I. Tesser,et al.  Synthesis, resolution and charge-donor properties of six tryptophan analogues. , 2009, International journal of peptide and protein research.

[16]  M. Saraste,et al.  FEBS Lett , 2000 .

[17]  N. Unwin.  Nicotinic acetylcholine receptor at 9 A resolution. , 1993, Journal of molecular biology.

[18]  Sandro Mecozzi,et al.  Cation−π Interactions in Simple Aromatics: Electrostatics Provide a Predictive Tool , 1996 .

[19]  A. Karlin,et al.  Acetylcholine Receptor: Covalent Attachment of Depolarizing Groups at the Active Site , 1969, Science.

[20]  N. Davidson,et al.  Nicotinic receptor binding site probed with unnatural amino acid incorporation in intact cells. , 1995, Science.

[21]  J. Changeux,et al.  Mutations in the channel domain alter desensitization of a neuronal nicotinic receptor , 1991, Nature.

[22]  P G Schultz,et al.  A general method for site-specific incorporation of unnatural amino acids into proteins. , 1989, Science.

[23]  Manuel Peitsch,et al.  Probing the Structure and Function of the Tachykinin Neurokinin-2 Receptor through Biosynthetic Incorporation of Fluorescent Amino Acids at Specific Sites* , 1996, The Journal of Biological Chemistry.

[24]  J. Galzi,et al.  Neuronal nicotinic receptors: Molecular organization and regulations , 1995, Neuropharmacology.

[25]  J. Changeux,et al.  Functional significance of aromatic amino acids from three peptide loops of the α7 neuronal nicotinic receptor site investigated by site‐directed mutagenesis , 1991, FEBS letters.

[26]  N S Scrutton,et al.  Cation-pi bonding and amino-aromatic interactions in the biomolecular recognition of substituted ammonium ligands. , 1996, The Biochemical journal.

[27]  P. Schultz,et al.  Site-directed mutagenesis with an expanded genetic code. , 1995, Annual review of biophysics and biomolecular structure.

[28]  Burton S. Rosner,et al.  Neuropharmacology , 1958, Nature.

[29]  Baldev Singh,et al.  Synthesis of 7-azaindole and 7-azaoxindole derivatives through a palladium-catalyzed cross-coupling reaction , 1992 .

[30]  M. M. White,et al.  Ligand—receptor interactions in the nicotinic acetylcholine receptor probed using multiple substitutions at conserved tyrosines on the α subunit , 1994, FEBS letters.