Universidad Nacional Autónoma de México, México DF 04510, México.
J Med Chem. 2011 Jun 9;54(11):3875-84. doi: 10.1021/jm200167g. Epub 2011 May 5.
A novel, sensible, and specific fluorescent biosensor of human calmodulin (hCaM), namely hCaM L39C-mBBr/V91C-mBBr, was constructed. The biosensor was useful for detecting ligands with opposing fluorescent signals, calcium ions (Ca(2+)) and CaM inhibitors in solution. Thus, the device was successfully applied to analyze the allosteric effect of Ca(2+) on trifluoroperazine (TFP) binding to CaM (Ca(2+)K(d) = 0.24 μM ± 0.03 with a stoichiometry 4.10 ± 0.15; TFPK(d) ∼ 5.74-0.53 μM depending on the degree of saturation of Ca(2+), with a stoichiometry of 2:1). In addition, it was suitable for discovering additional xanthones (5, 6, and 8) with anti-CaM properties from the fungus Emericella 25379. The affinity of 1-5, 7, and 8 for the complex (Ca(2+))(4)-CaM was excellent because their experimental K(d)s were in the nM range (4-498 nM). Docking analysis predicted that 1-8 bind to CaM at sites I, III, and IV as does TFP.
构建了一种新型的、灵敏的、特异性的人钙调蛋白（hCaM）荧光生物传感器，即 hCaM L39C-mBBr/V91C-mBBr。该生物传感器可用于检测溶液中具有相反荧光信号的配体、钙离子（Ca(2+)）和 CaM 抑制剂。因此，该装置成功地应用于分析 Ca(2+)对三氟拉嗪（TFP）与 CaM 结合的变构效应（Ca(2+)K(d) = 0.24 μM ± 0.03，化学计量比为 4.10 ± 0.15；TFPK(d)∼5.74-0.53 μM，取决于 Ca(2+)的饱和度，化学计量比为 2:1）。此外，它还适用于从真菌 Emericella 25379 中发现具有抗 CaM 特性的其他紫檀烷（5、6 和 8）。由于它们的实验 K(d) 值在纳摩尔范围内（4-498 nM），因此 1-5、7 和 8 对复合物（Ca(2+))(4)-CaM 的亲和力非常好。对接分析预测 1-8 与 TFP 一样，与 CaM 的结合部位在 I、III 和 IV 上。
