Xiu X et al. (2009), Nicotine binding to brain receptors requires a ...

XB-ART-39704

Nature 2009 Mar 26;4587237:534-7. doi: 10.1038/nature07768.

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Nicotine binding to brain receptors requires a strong cation-pi interaction.

Xiu X , Puskar NL , Shanata JA , Lester HA , Dougherty DA .

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Nicotine addiction begins with high-affinity binding of nicotine to acetylcholine (ACh) receptors in the brain. The end result is over 4,000,000 smoking-related deaths annually worldwide and the largest source of preventable mortality in developed countries. Stress reduction, pleasure, improved cognition and other central nervous system effects are strongly associated with smoking. However, if nicotine activated ACh receptors found in muscle as potently as it does brain ACh receptors, smoking would cause intolerable and perhaps fatal muscle contractions. Despite extensive pharmacological, functional and structural studies of ACh receptors, the basis for the differential action of nicotine on brain compared with muscle ACh receptors has not been determined. Here we show that at the alpha4beta2 brain receptors thought to underlie nicotine addiction, the high affinity for nicotine is the result of a strong cation-pi interaction to a specific aromatic amino acid of the receptor, TrpB. In contrast, the low affinity for nicotine at the muscle-type ACh receptor is largely due to the fact that this key interaction is absent, even though the immediate binding site residues, including the key amino acid TrpB, are identical in the brain and muscle receptors. At the same time a hydrogen bond from nicotine to the backbone carbonyl of TrpB is enhanced in the neuronal receptor relative to the muscle type. A point mutation near TrpB that differentiates alpha4beta2 and muscle-type receptors seems to influence the shape of the binding site, allowing nicotine to interact more strongly with TrpB in the neuronal receptor. ACh receptors are established therapeutic targets for Alzheimer's disease, schizophrenia, Parkinson's disease, smoking cessation, pain, attention-deficit hyperactivity disorder, epilepsy, autism and depression. Along with solving a chemical mystery in nicotine addiction, our results provide guidance for efforts to develop drugs that target specific types of nicotinic receptors.

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Species referenced: Xenopus laevis
Genes referenced: cntn1

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	Figure 2. Agonists and unnatural amino acids considered here. a. Structures of ACh and nicotine. b. Unnatural amino acids considered here. If not indicated, an a, b, c, or d group is H. c. The backbone ester strategy for modulating a hydrogen bond.
	Figure 3. Nonsense suppression in the Î±4Î²2 receptor. Shown is a wild type recovery experiment, in which Trp is incorporated at the TrpB position. a. Representative traces of voltage-clamp currents. Bars represent application of ACh at concentrations noted. b. Fit of data in A to the Hill equation.
	Figure 4. Fluorination plots. Note that in both plots, all data sets share the point at x = 32.6 kcal/mol (cation-Ï energy for Trp); y = 0 (black circle). Moving to the left then corresponds to monofluoro-, difluoro-, trifluoro- and tetrafluoro-TrpB. Cation-Ï binding energies (x-axes) are from 9. a. Muscle-type receptor. The designation âwt" indicates G at position 153. b. Î±4Î²2 receptor.
	Figure 5. Single-channel recordings from wild type Î±4Î²2 (conventional expression) and the F3-Trp mutant (nonsense suppression) at site B, with nicotine applied at EC50 values (0.080 and 1.2 ÂµM, respectively). Lower traces are expansions of the regions marked by a bar in the upper trace. Records were obtained in the cell-attached configuration with a pipette potential of +100 mV and are shown at 2 kHz bandwidth. Channel openings are shown as downward deflections.