Physical Chemistry and Mass Spectrometry Laboratory, Department of Chemistry, Building B6c, University of Liège, B-4000 Liège, Belgium.
Anal Chem. 2009 Aug 15;81(16):6708-15. doi: 10.1021/ac900785m.
The quantitative study of noncovalent complexes by electrospray mass spectrometry requires the determination of the relative response of each species. The method proposed here to determine the electrospray response factors is based on the use of (1) an internal standard and (2) the mass balance equation applied to one binding partner M, for which different complexes M(x)L(y) are detected in the electrospray mass spectra. A set of experiments providing various ratios between the complexes (e.g., different ligand concentrations in a titration experiment or different time points in a kinetics experiment) is used to generate a set of independent linear equations that can be solved using simple matrix algebra to find the response factors of each M(x)L(y) complex relative to that of the internal standard. The response factors can then be used to determine equilibrium dissociation constants or for the quantitative monitoring of reaction kinetics. The first is illustrated with a study of DNA-ligand complexes, where we show that neither minor groove binding nor intercalation dramatically affects the DNA response factor. The second is illustrated with a study of the association kinetics of the telomeric G-quadruplex dGGG(TTAGGG)(3) with its complementary strand, where the response factors allow correcting for the relative response of the quadruplex and the long duplex and obtaining reproducible association rate constants independently of the source tuning potentials.
通过电喷雾质谱法对非共价复合物进行定量研究需要确定每种物质的相对响应。这里提出的确定电喷雾响应因子的方法基于使用（1）内标和（2）应用于一个结合配偶体 M 的质量平衡方程，对于 M，可以在电喷雾质谱中检测到不同的复合物 M(x)L(y)。一组提供复合物之间各种比例的实验（例如，滴定实验中的不同配体浓度或动力学实验中的不同时间点）用于生成一组独立的线性方程，可以使用简单的矩阵代数来求解这些方程，以找到每个 M(x)L(y)复合物相对于内标物的响应因子。然后可以使用响应因子来确定平衡离解常数或定量监测反应动力学。第一个例子是 DNA-配体复合物的研究，我们表明，无论是小沟结合还是嵌入都不会显著影响 DNA 的响应因子。第二个例子是端粒 G-四联体 dGGG(TTAGGG)(3)与其互补链的缔合动力学研究，其中响应因子允许校正四联体和长双链的相对响应，并获得可重现的缔合速率常数，而与源调谐电位无关。
