An explicit formulation approach for the analysis of calcium binding to EF-hand proteins using isothermal titration calorimetry.
An explicit formulation approach for the analysis of calcium binding to EF-hand proteins using isothermal titration calorimetry.
复制标题
使用等温滴定量热法分析钙与 EF-hand 蛋白结合的明确配方方法。
DOI：
10.1016/j.bpj.2013.11.017
复制
发表时间：
2013
影响因子：
3.4
通讯作者：
Hodsdon,MichaelE
中科院分区：
文献类型：
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作者：
Keeler,Camille;Poon,Gregory;Kuo,IvanaY;Ehrlich,BarbaraE;Hodsdon,MichaelE
我们提出了一种改进和扩展的数学方法，用于从等温滴定量热法模拟配体与大分子的结合等温线。我们的方法使用作为初值问题的隐式和数值求解的常微分方程组，从一般结合多项式的基础上提供对复杂的大分子混合物中每个竞争成员的分数界限的定量描述。这种方法极大地简化了复杂结合模型的公式。除了我们通用的无模型方法外，我们还引入了一种数学处理方法，用于处理在滴定开始之前配体存在的情况，当完全移除结合伙伴可能会破坏大分子的结构和功能特征时，这对于数据分析是必不可少的。提供了可在免费获得的软件平台上播放的演示程序。我们的方法用经典的钙离子选择电位法和钙离子与混合络合剂的结合等温线进行了实验验证，反应容器中存在和不存在残留配体。最后，我们模拟并比较了与多囊肾病相关的钙依赖通道多囊蛋白2的钙结合等温线与其典型结合部位(EF-HAND结构域)的结合。
We present an improved and extended version of a recently proposed mathematical approach for modeling isotherms of ligand-to-macromolecule binding from isothermal titration calorimetry. Our approach uses ordinary differential equations, solved implicitly and numerically as initial value problems, to provide a quantitative description of the fraction bound of each competing member of a complex mixture of macromolecules from the basis of general binding polynomials. This approach greatly simplifies the formulation of complex binding models. In addition to our generalized, model-free approach, we have introduced a mathematical treatment for the case where ligand is present before the onset of the titration, essential for data analysis when complete removal of the binding partner may disrupt the structural and functional characteristics of the macromolecule. Demonstration programs playable on a freely available software platform are provided. Our method is experimentally validated with classic calcium (Ca2+) ion-selective potentiometry and isotherms of Ca2+binding to a mixture of chelators with and without residual ligand present in the reaction vessel. Finally, we simulate and compare experimental data fits for the binding isotherms of Ca2+binding to its canonical binding site (EF-hand domain) of polycystin 2, a Ca2+-dependent channel with relevance to polycystic kidney disease.