TY - JOUR
T1 - Molecular determinants of subtype-selective efficacies of cytisine and the novel compound NS3861 at heteromeric nicotinic acetylcholine receptors
AU - Harpsøe, Kasper
AU - Hald, Helle
AU - Timmermann, Daniel B
AU - Jensen, Marianne L
AU - Dyhring, Tino
AU - Nielsen, Elsebet Ø
AU - Peters, Dan
AU - Balle, Thomas
AU - Gajhede, Michael
AU - Kastrup, Jette Sandholm Jensen
AU - Ahring, Philip K
PY - 2013/1/25
Y1 - 2013/1/25
N2 - Deciphering which specific agonist-receptor interactions affect efficacy levels is of high importance, because this will ultimately aid in designing selective drugs. The novel compound NS3861 and cytisine are agonists of nicotinic acetylcholine receptors (nAChRs) and both bind with high affinity to heteromeric α3β4 and α4β2 nAChRs. However, initial data revealed that the activation patterns of the two compounds show very distinct maximal efficacy readouts at various heteromeric nAChRs. To investigate the molecular determinants behind these observations, we performed in-depth patch clamp electrophysiological measurements of efficacy levels at heteromeric combinations of α3- and α4-, with β2- and β4-subunits, and various chimeric constructs thereof. Compared with cytisine, which selectively activates receptors containing β4- but not β2-subunits, NS3861 displays the opposite β-subunit preference and a complete lack of activation at α4-containing receptors. The maximal efficacy of NS3861 appeared solely dependent on the nature of the ligand-binding domain, whereas efficacy of cytisine was additionally affected by the nature of the β-subunit transmembrane domain. Molecular docking to nAChR subtype homology models suggests agonist specific interactions to two different residues on the complementary subunits as responsible for the β-subunit preference of both compounds. Furthermore, a principal subunit serine to threonine substitution may explain the lack of NS3861 activation at α4-containing receptors. In conclusion, our results are consistent with a hypothesis where agonist interactions with the principal subunit (α) primarily determine binding affinity, whereas interactions with key amino acids at the complementary subunit (β) affect agonist efficacy.
AB - Deciphering which specific agonist-receptor interactions affect efficacy levels is of high importance, because this will ultimately aid in designing selective drugs. The novel compound NS3861 and cytisine are agonists of nicotinic acetylcholine receptors (nAChRs) and both bind with high affinity to heteromeric α3β4 and α4β2 nAChRs. However, initial data revealed that the activation patterns of the two compounds show very distinct maximal efficacy readouts at various heteromeric nAChRs. To investigate the molecular determinants behind these observations, we performed in-depth patch clamp electrophysiological measurements of efficacy levels at heteromeric combinations of α3- and α4-, with β2- and β4-subunits, and various chimeric constructs thereof. Compared with cytisine, which selectively activates receptors containing β4- but not β2-subunits, NS3861 displays the opposite β-subunit preference and a complete lack of activation at α4-containing receptors. The maximal efficacy of NS3861 appeared solely dependent on the nature of the ligand-binding domain, whereas efficacy of cytisine was additionally affected by the nature of the β-subunit transmembrane domain. Molecular docking to nAChR subtype homology models suggests agonist specific interactions to two different residues on the complementary subunits as responsible for the β-subunit preference of both compounds. Furthermore, a principal subunit serine to threonine substitution may explain the lack of NS3861 activation at α4-containing receptors. In conclusion, our results are consistent with a hypothesis where agonist interactions with the principal subunit (α) primarily determine binding affinity, whereas interactions with key amino acids at the complementary subunit (β) affect agonist efficacy.
U2 - 10.1074/jbc.m112.436337
DO - 10.1074/jbc.m112.436337
M3 - Journal article
C2 - 23229547
SN - 0021-9258
VL - 288
SP - 2559
EP - 2570
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 4
ER -