Calcium-induced changes in calmodulin structural dynamics and thermodynamics

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• 作者：Guangrong Wu^a ; Zhengya Gao^a ; Aichun Dong^b ; ^{Aichun.Dong@unco.edu} ; Shaoning Yu^a ; ^{yushaoning@fudan.edu.cn}
• 关键词：CaM ; calmodulin ; ITC ; isothermal titration calorimetry ; FT-IR ; Fourier transform infrared ; IPTG ; isopropylthiogalactopyranoside ; CD ; circular dichroism ; GdnHCl ; guanidine hydrochloride ; H&ndash ; D exchange ; hydrogen&ndash ; deuterium exchange
• 刊名：International Journal of Biological Macromolecules
• 出版年：2012
• 期刊代码：62_01418130
• 类别：abs
• 出版时间：1 May, 2012
• 卷：50
• 期：4
• 页码：1011-1017
• 文件大小：623 K

摘要

The thermodynamics of the interaction between Ca²⁺ and calmodulin (CaM) was examined using isothermal titration calorimetry (ITC). The chemical denaturation of calmodulin was monitored spectroscopically to determine the stability of Ca²⁺-free (apo) and Ca²⁺-loaded (holo) CaMs. We explored the conformational and structural dynamics of CaM using amide hydrogen-deuterium (H-D) exchange coupled with Fourier transform infrared (FT-IR) spectroscopy. The results of H-D exchange and FT-IR suggest that CaM activation by Ca²⁺ binding involves significant conformational changes. The results have also revealed that while the overall conformation of holo-CaM is more stable than that of the apo-CaM, some part of its 伪-helix structures, most likely the EF-hand domain region, has more solvent exposure, thus, has a faster H-D exchange rate than that of the apo-CaM. The ITC method provides a new strategy for obtaining site-specific Ca²⁺ binding properties and a better estimation of the cooperativity and conformational change contributions of coupled EF-hand proteins.