Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Varenicline Interactions at the 5-HT3 Receptor Ligand Binding Site are Revealed by 5-HTBP.
Price KL
,
Lillestol RK
,
Ulens C
,
Lummis SC
.
???displayArticle.abstract???
Cys-loop receptors are the site of action of many therapeutic drugs. One of these is the smoking cessation agent varenicline, which has its major therapeutic effects at nicotinic acetylcholine (nACh) receptors but also acts at 5-HT3 receptors. Here, we report the X-ray crystal structure of the 5-HT binding protein (5-HTBP) in complex with varenicline, and test the predicted interactions by probing the potency of varenicline in a range of mutant 5-HT3 receptors expressed in HEK293 cells and Xenopus oocytes. The structure reveals a range of interactions between varenicline and 5-HTBP. We identified residues within 5 Å of varenicline and substituted the equivalent residues in the 5-HT3 receptor with Ala or a residue with similar chemical properties. Functional characterization of these mutant 5-HT3 receptors, using a fluorescent membrane potential dye in HEK cells and voltage clamp in oocytes, supports interactions between varenicline and the receptor that are similar to those in 5-HTBP. The structure also revealed C-loop closure that was less than in the 5-HT-bound 5-HTBP, and hydrogen bonding between varenicline and the complementary face of the binding pocket via a water molecule, which are characteristics consistent with partial agonist behavior of varenicline in the 5-HT3 receptor. Together, these data reveal detailed insights into the molecular interaction of varenicline in the 5-HT3 receptor.
Figure 1. Varenicline bound to
5-HTBP. (A) Location of varenicline (green)
at the interface between two subunits in the orthosteric 5-HTBP binding
site. (B) Alignment of 5-HTBP, AChBP, and the extracellular domain
of the 5-HT3A receptor subunit showing the approximate
location of the AâE binding loops. The residues mutated in
this study are highlighted in purple, and the residues that differ
between 5-HTBP and AChBP in yellow. (C) The 5-HTBP binding pocket
showing the orientation of varenicline (green) and nearby residues
on the principal (blue) and complementary (magenta) faces. The corresponding
5-HT3 receptor residues are in parentheses. (D) Hydrogen
bonds are present between varenicline and residues Y91, Y193, and
W145 on the principal face and residues I104 and I116 on the complementary
face via a water molecule.
Figure 2. HEK cell data. (A) Typical traces from HEK cells transfected with
WT 5-HT3 receptor cDNA, loaded with membrane potential
dye, and stimulated at 20 s with various concentrations of varenicline. F = arbitrary fluorescent units. (B) Concentrationâresponse
curves constructed from FlexStation responses to 5-HT (squares, filled
line) and varenicline (circles, dashed line). Data = mean ± SEM, n = 4.
Figure 3. Concentration
response curves for WT and mutant 5-HT3 receptors expressed
in oocytes. Concentration response curves for
(A) 5-HT3A L184A and (B) 5-HT3A W90Y mutant
receptors (gray lines) compared to WT 5-HT3A receptors
(black lines) for 5-HT (solid lines) and varenicline (dashed lines)
reveal differences in EC50 and Rmax/Rmax,5-HT. Data = mean ±
SEM, n = 3â6.
Figure 4. The 5-HT3 receptor loop E region. Structural data from
the mouse 5-HT3A receptor structure (PDB: 4PIR) reveal a network
of hydrogen bonds involving the loop E aromatic residues Y141, Y143,
and Y153. Functional data show the importance of these residues for
proper functioning of the receptor.
Beene,
Cation-pi interactions in ligand recognition by serotonergic (5-HT3A) and nicotinic acetylcholine receptors: the anomalous binding properties of nicotine.
2002, Pubmed,
Xenbase
Beene,
Cation-pi interactions in ligand recognition by serotonergic (5-HT3A) and nicotinic acetylcholine receptors: the anomalous binding properties of nicotine.
2002,
Pubmed
,
Xenbase
Beene,
Tyrosine residues that control binding and gating in the 5-hydroxytryptamine3 receptor revealed by unnatural amino acid mutagenesis.
2004,
Pubmed
,
Xenbase
Billen,
Molecular actions of smoking cessation drugs at α4β2 nicotinic receptors defined in crystal structures of a homologous binding protein.
2012,
Pubmed
,
Xenbase
Blum,
Nicotinic pharmacophore: the pyridine N of nicotine and carbonyl of acetylcholine hydrogen bond across a subunit interface to a backbone NH.
2010,
Pubmed
,
Xenbase
Brams,
A structural and mutagenic blueprint for molecular recognition of strychnine and d-tubocurarine by different cys-loop receptors.
2011,
Pubmed
Fagerström,
Varenicline in the treatment of tobacco dependence.
2008,
Pubmed
Gurley,
Nicotinic agonists competitively antagonize serotonin at mouse 5-HT3 receptors expressed in Xenopus oocytes.
1998,
Pubmed
,
Xenbase
Hansen,
Structures of Aplysia AChBP complexes with nicotinic agonists and antagonists reveal distinctive binding interfaces and conformations.
2005,
Pubmed
Hassaine,
X-ray structure of the mouse serotonin 5-HT3 receptor.
2014,
Pubmed
Hibbs,
Structural determinants for interaction of partial agonists with acetylcholine binding protein and neuronal alpha7 nicotinic acetylcholine receptor.
2009,
Pubmed
Holbrook,
Characterisation of 5-HT3C, 5-HT3D and 5-HT3E receptor subunits: evolution, distribution and function.
2009,
Pubmed
Joshi,
Interactions of granisetron with an agonist-free 5-HT3A receptor model.
2006,
Pubmed
,
Xenbase
Kesters,
Structural basis of ligand recognition in 5-HT3 receptors.
2013,
Pubmed
Lummis,
Varenicline is a potent agonist of the human 5-hydroxytryptamine3 receptor.
2011,
Pubmed
,
Xenbase
Lummis,
5-HT(3) receptors.
2012,
Pubmed
Machu,
Therapeutics of 5-HT3 receptor antagonists: current uses and future directions.
2011,
Pubmed
Mihalak,
Varenicline is a partial agonist at alpha4beta2 and a full agonist at alpha7 neuronal nicotinic receptors.
2006,
Pubmed
,
Xenbase
Miles,
A coupled array of noncovalent interactions impacts the function of the 5-HT3A serotonin receptor in an agonist-specific way.
2012,
Pubmed
,
Xenbase
Niesler,
Characterization of the novel human serotonin receptor subunits 5-HT3C,5-HT3D, and 5-HT3E.
2007,
Pubmed
Price,
A hydrogen bond in loop A is critical for the binding and function of the 5-HT3 receptor.
2008,
Pubmed
Price,
FlexStation examination of 5-HT3 receptor function using Ca2+ - and membrane potential-sensitive dyes: advantages and potential problems.
2005,
Pubmed
Puskar,
Two neuronal nicotinic acetylcholine receptors, alpha4beta4 and alpha7, show differential agonist binding modes.
2011,
Pubmed
,
Xenbase
Rohde,
Intersubunit bridge formation governs agonist efficacy at nicotinic acetylcholine α4β2 receptors: unique role of halogen bonding revealed.
2012,
Pubmed
Rucktooa,
Structural characterization of binding mode of smoking cessation drugs to nicotinic acetylcholine receptors through study of ligand complexes with acetylcholine-binding protein.
2012,
Pubmed
Spier,
The role of tryptophan residues in the 5-Hydroxytryptamine(3) receptor ligand binding domain.
2000,
Pubmed
Tavares,
Variations in binding among several agonists at two stoichiometries of the neuronal, α4β2 nicotinic receptor.
2012,
Pubmed
Thompson,
5-HT3 receptors.
2006,
Pubmed
Thompson,
Loop B is a major structural component of the 5-HT3 receptor.
2008,
Pubmed
Thompson,
Locating an antagonist in the 5-HT3 receptor binding site using modeling and radioligand binding.
2005,
Pubmed
Van Arnam,
An unusual pattern of ligand-receptor interactions for the α7 nicotinic acetylcholine receptor, with implications for the binding of varenicline.
2013,
Pubmed
,
Xenbase
