Anal Chem. 2010 Mar 15;82(6):2170-4. doi: 10.1021/ac902633d.
The application of the direct electrospray ionization mass spectrometry (ES-MS) assay for quantifying the stoichiometry and absolute affinity of protein-metal ion binding in vitro is described. Control ES-MS experiments performed on solutions containing calcium chloride or calcium acetate and a pair of proteins that do not bind calcium ions in solution revealed that the nonspecific association of metal ions to proteins during ES is a random process, independent of protein size and structure. These results establish the reliability of the reference protein method for quantitatively correcting ES mass spectra for the occurrence of nonspecific metal ion binding to proteins during ES-MS analysis. To demonstrate the utility of the direct ES-MS assay, when carried out using the reference protein method, the calcium binding stoichiometry of bovine alpha-lactalbumin and the calcium ion affinity of bovine beta-lactoglobulin were established.
本文描述了直接电喷雾电离质谱（ESI-MS）测定在体外定量蛋白质-金属离子结合的化学计量和绝对亲和力的应用。在含有氯化钙或醋酸钙以及一对在溶液中不结合钙离子的蛋白质的溶液上进行的对照 ESI-MS 实验表明，ESI 过程中金属离子与蛋白质的非特异性结合是一个随机过程，与蛋白质的大小和结构无关。这些结果证实了参考蛋白法的可靠性，可用于定量校正 ESI-MS 谱中非特异性金属离子与蛋白质结合的发生，以进行 ESI-MS 分析。为了证明直接 ESI-MS 测定法的实用性，当使用参考蛋白法进行时，确定了牛α-乳白蛋白的钙结合化学计量和牛β-乳球蛋白的钙离子亲和力。
