Haikala H, Kaivola J, Nissinen E, Wall P, Levijoki J, Lindén I B
Orion-Farmos, Orion Research Center, Espoo, Finland.
J Mol Cell Cardiol. 1995 Sep;27(9):1859-66. doi: 10.1016/0022-2828(95)90009-8.
The role of cardiac troponin C (cTnC) as a target protein for the calcium sensitization by levosimendan, pimobendan, MCI-154 and EMD 53998 was evaluated using purified recombinant human cTnC. For determination of calcium- and magnesium-dependent binding of the compounds to cTnC a new type of cTnC-HPLAC column was used. Furthermore, dansylated cTnC was utilized to study the effect of the calcium sensitizing compounds on calcium-induced conformation of cTnC. Only levosimendan showed calcium-dependent and to a lesser extent magnesium-dependent retention in the cTnC column. The findings indicate that levosimendan binds both to the N-terminal and C-terminal domains of cTnC. In agreement with this, only levosimendan shifted the calcium-induced fluorescence curve of dansylated cTnC to the left. In the control experiments Ca50 and KCa2+ were calculated to be 2.73 microM and 4 x 10(5) M-1, respectively. Levosimendan at 3 microM decreased the value of Ca50 to 1.19 microM. In conclusion, it is suggested that the mechanism of calcium sensitizing effect of levosimendan, unlike that of the other calcium sensitizers, is based on calcium-dependent binding to the N-terminal domain of cTnC. This is proposed to amplify the trigger of contraction induced by cTnC in the cardiac muscle.
利用纯化的重组人心肌肌钙蛋白C（cTnC）评估了左西孟旦、匹莫苯丹、MCI-154和EMD 53998作为钙增敏作用靶蛋白的作用。为了测定这些化合物与cTnC的钙和镁依赖性结合，使用了一种新型的cTnC-HPLAC柱。此外，利用丹磺酰化的cTnC研究钙增敏化合物对钙诱导的cTnC构象的影响。只有左西孟旦在cTnC柱中表现出钙依赖性保留，在较小程度上表现出镁依赖性保留。研究结果表明，左西孟旦与cTnC的N端和C端结构域均有结合。与此一致的是，只有左西孟旦将丹磺酰化cTnC的钙诱导荧光曲线向左移动。在对照实验中，计算得出Ca50和KCa2+分别为2.73 microM和4×10(5) M-1。3 microM的左西孟旦将Ca50值降低至1.19 microM。总之，有人提出，与其他钙增敏剂不同，左西孟旦的钙增敏作用机制是基于其与cTnC N端结构域的钙依赖性结合。这被认为可放大心肌中cTnC诱导的收缩触发作用。
