ANALYSIS OF THE ION-BINDING SITES OF CALMODULIN BY ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY
ANALYSIS OF THE ION-BINDING SITES OF CALMODULIN BY ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY
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DOI：
10.1021/bi00042a014
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发表时间：
1995-10-24
期刊：
影响因子：
2.9
通讯作者：
CALAS, B
中科院分区：
文献类型：
--
作者：
LAFITTE, D;CAPONY, JP;CALAS, B
通过 ESI-MS 研究了 Ca2+ 和 Mg2+ 与四种钙调蛋白（SynCaM 1、SynCaM 8、SynCaM 12A 和 SynCaM 18A）的结合。通过将脱辅基蛋白溶解在含有 1 mM CaCl2 或 1 mM MgCl2 的甲醇/水（20/80，v/v）中并用氨将 pH 调节至 6.0 来记录质谱。载体溶剂是甲醇/水(20/80，v/v)。对于 Ca2+ 络合，ESI-MS 显示存在三种位点：第一种位点具有高亲和力，对应于使用流动和平衡透析技术确定的位点，另外两种位点具有较低的亲和力。这些结果清楚地证实了 Miles 等人的结论。 [Miles, M.、Comte, M.、Schaer, J. J. 和 Cox, J. A. (1989) J. Inorg。生物化学。 36, 11-25]应该存在四到六个Ca2+辅助位点。关于镁的络合，四种蛋白质几乎肯定能够在辅助阳离子位点上结合两个 Mg2+。
The binding of Ca2+ and Mg2+ to four calmodulins (SynCaM 1, SynCaM 8, SynCaM 12A, and SynCaM 18A) has been studied by ESI-MS. The mass spectra were recorded by dissolving the apoproteins in methanol/water (20/80, v/v) containing 1 mM CaCl2 or 1 mM MgCl2 and the pH adjusted to 6.0 with ammonia. The carrier solvent was methanol/water (20/80, v/v). In the case of Ca2+ complexation, ESI-MS reveals the presence of three kinds of sites: the first of high affinity corresponding to those determined using flow and equilibrium dialysis techniques and two others with lower affinities. These results clearly confirm the conclusion of Miles et al. [Miles, M., Comte, M., Schaer, J. J., & Cox, J. A. (1989) J. Inorg. Biochem. 36, 11-25] that there should exist between four and six auxiliary sites for Ca2+. Concerning the complexation of magnesium, the four proteins are able to bind two Mg2+ almost certainly on auxiliary cationic sites.