Structure, dynamics, and thermodynamics of the structural domain of troponin C in complex with the regulatory peptide 1-40 of troponin I
Structure, dynamics, and thermodynamics of the structural domain of troponin C in complex with the regulatory peptide 1-40 of troponin I
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DOI：
10.1021/bi010748
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发表时间：
2001-08-28
期刊：
影响因子：
2.9
通讯作者：
Sykes, BD
中科院分区：
文献类型：
--
作者：
Mercier, P;Spyracopoulos, L;Sykes, BD
采用核磁共振（NMR）技术测定了由肌钙蛋白I （Tnl）残基1-40 （Rp40）组成的调节肽复合物中骨骼肌钙蛋白C （CTnC）钙饱和C结构域的结构。核磁共振测定的溶液结构与x射线晶体学测定的完整TnC与TnI（1-47）复合物的c域结构相似[Vassylyev， D. G., Takeda， S., Wakatsuki， S., Maeda， K., and Maeda， Y. (1998) Proc. Nad.]。学会科学。[j]。CTnC动态特性的变化。利用主酰胺N-15核磁共振弛豫测量方法研究了Rp40结合诱导的2Ca（2+）。对核磁共振弛豫数据的分析允许在每个残基的基础上提取运动顺序参数，从中可以估计皮秒到纳秒时间尺度运动对与复杂形成相关的构象熵的贡献。结果表明，Rp40的结合降低了CTnC中主干的柔韧性，特别是在c末端螺旋的末端。主链构象熵变化（-T δ）与Rp40与CTnC的结合有关。根据N-15弛豫数据测定的2Ca（2+）在30°c时为9.6 +/- 0.7 kcal mol（-1）。然而，使用结构方法估算热力学量[Lavigne， P., Bagu， J. R., Boyko， R., Willard， L., Holmes， C. F.和Sykes， B. D. (2000) Protein Sci. 9,252 -264]表明，复合物形成时溶剂化熵的变化占主导地位，并且克服了与Rp40结合时蛋白质骨架“硬化”相关的热力学“成本”。此外，在不同浓度的CTnC下测量了骨干酰胺15N弛豫数据。2 ca(2 +)。Rp40表明该配合物在溶液中二聚体化。将表观全球旋转相关时间作为浓度的函数拟合到单体-二聚体平衡中，得到二聚常数近似于8.3 mM。
The structure of the calcium-saturated C-domain of skeletal troponin C (CTnC) in complex with a regulatory peptide comprising residues 1-40 (Rp40) of troponin I (Tnl) was determined using nuclear magnetic resonance (NMR) spectroscopy. The solution structure determined by NMR is similar to the structure of the C-domain from intact TnC in complex with TnI(1-47) determined by X-ray crystallography [Vassylyev, D. G., Takeda, S., Wakatsuki, S., Maeda, K., and Maeda, Y. (1998) Proc. Nad. Acad. Sci. U.S.A. 95, 4847-4852]. Changes in the dynamic properties of CTnC . 2Ca(2+) induced by Rp40 binding were investigated using backbone amide N-15 NMR relaxation measurements. Analysis of NMR relaxation data allows for extraction of motional order parameters on a per residue basis, from which the contribution of changes in picosecond to nanosecond time scale motions to the conformational entropy associated with complex formation can be estimated. The results indicate that binding of Rp40 decreases backbone flexibility in CTnC, particularly at the end of the C-terminal helix. The backbone conformational entropy change (-T DeltaS) associated with binding of Rp40 to CTnC . 2Ca(2+) determined from N-15 relaxation data is 9.6 +/- 0.7 kcal mol(-1) at 30 degreesC. However, estimation of thermodynamic quantities using a structural approach [Lavigne, P., Bagu, J. R., Boyko, R., Willard, L., Holmes, C. F., and Sykes, B. D. (2000) Protein Sci. 9, 252-264] reveals that the change in solvation entropy upon complex formation is dominant and overcomes the thermodynamic "cost" associated with "stiffening" of the protein backbone upon Rp40 binding. Additionally, backbone amide 15N relaxation data measured at different concentrations of CTnC . 2Ca(2+). Rp40 reveal that the complex dimerizes in solution. Fitting of the apparent global rotational correlation time as a function of concentration to a monomer-dimer equilibrium yields a dimerization constant of similar to8.3 mM.