TY - JOUR T1 - Key Roles of Phe<sup>1112</sup> and Ser<sup>1115</sup> in the Pore-Forming IIIS5-S6 Linker of L-Type Ca<sup>2+</sup> Channel α<sub>1C</sub> Subunit (Ca<sub>V</sub> 1.2) in Binding of Dihydropyridines and Action of Ca<sup>2+</sup> Channel Agonists JF - Molecular Pharmacology JO - Mol Pharmacol SP - 235 LP - 248 DO - 10.1124/mol.64.2.235 VL - 64 IS - 2 AU - Shinji Yamaguchi AU - Boris S. Zhorov AU - Katsuro Yoshioka AU - Taku Nagao AU - Hidenori Ichijo AU - Satomi Adachi-Akahane Y1 - 2003/08/01 UR - http://molpharm.aspetjournals.org/content/64/2/235.abstract N2 - Voltage-dependent L-type Ca2+ channels are modulated by the binding of Ca2+ channel antagonists and agonists to the pore-forming α1c subunit (CaV 1.2). We recently identified Ser1115 in IIIS5-S6 linker of α1C subunit as a critical determinant of the action of 1,4-dihydropyridine agonists. In this study, we applied alanine-scanning mutational analysis in IIIS5-S6 linker of rat brain α1C subunit (rbCII) to illustrate the role of pore-forming IIIS5-S6 linker in the action of Ca2+ channel modulators. Ca2+ channel currents through wild-type (rbCII) or mutated α1C subunits, transiently expressed in BHK6 cells with β1a and α2/δ subunits, were analyzed. The replacement of Phe1112 by Ala (F1112A) significantly impaired the sensitivity to Ca2+ channel agonists (S)-(-)-Bay k 8644 and FPL-64176, and modestly to 1,4-dihydropyridine (DHP) antagonists. The low sensitivity of F1112A and S1115A to DHP antagonists was consistent with the reduced binding affinity for [3H](+)PN200-110. The replacement of Phe1112 by Tyr, but not by Ala, restored the long openings produced by FPL-64176, thus indicating the critical role of aromatic ring of Phe1112 in the Ca2+ channel agonist action. Interestingly, double-mutant Ca2+ channel (F1112A/S1115A) failed to discriminate between Ca2+ channel agonist (S)-(-)-1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-[trifluoromethyl] phenyl)-3-pyridine carboxylic acid methyl ester (Bay k 8644) and antagonist (R)-(+)-Bay k 8644 and was blocked by the two enantiomers in an identical manner. These results indicate that both Phe1112 and Ser1115 in linker IIIS5-S6 are required for the action of Ca2+ channel agonists. A model of the DHP receptor is proposed to visualize possible interactions of Phe1112, Ser1115, and other DHP-sensing residues with a typical DHP ligand nifedipine. ER -