NMR solution structure of calcium-saturated skeletal muscle troponin C.
NMR solution structure of calcium-saturated skeletal muscle troponin C.
复制标题
钙饱和骨骼肌肌钙蛋白 C 的 NMR 溶液结构。
作者：
Carolyn M. Slupsky;B. D. Sykes
肌钙蛋白 C (TnC) 是一种 18 kDa（162 个残基）的细丝钙结合蛋白，负责在肌浆网释放钙时触发肌肉收缩。结合两个钙离子的 TnC 结构先前已通过 X 射线方法解析。此处显示的是 TnC 的溶液结构，已使用 3D 和 4D 异核核磁共振 (NMR) 波谱技术解析。 1H、13C 和 15N 主链化学位移已发表 [Slupsky, C. M., Reinach, F. C., Smillie, L. B., & Sykes, B. D. (1995) Protein Sci. 4、1279-1290]。本文展示的是 1H、13C 和 15N 侧链化学位移，已完成 80%。钙饱和 TnC 的结构是根据 2106 NOE 衍生距离约束、121 phi 二面角约束和 76 psi 二面角约束确定的。钙饱和 TnC 的外观揭示了一个哑铃形分子，具有两个通过连接体连接的球状结构域。 N端和C端结构域的结构高度收敛[残基10-80和98-155的平均原子坐标位置的主链原子均方根偏差(rmsd)分别为0.66+/-0.17和0.69+/-0.18A]；然而，一个域相对于另一个域的方向没有明确定义，因此每个域在结构上似乎是独立的。本文测定的 TnC 的钙饱和形式与通过 X 射线方法测定的半饱和形式的比较揭示了两个主要差异。首先，N 端结构域发生重大结构变化，导致疏水袋打开，可能将其呈现给其靶蛋白肌钙蛋白 I。这种结构变化似乎仅涉及螺旋 B 和 C，它们通过主链 phi、psi 角的改变而远离螺旋 N/A/D 谷氨酸 41 从晶体结构中的不规则（-97 度，-7 度）到 NMR 钙饱和结构中的螺旋（-60度，-34 度）。两种结构之间的另一个区别是 NMR 结构中两个结构域之间存在灵活的连接体。这种灵活的接头允许 TnC 的两个结构域相互采用任何方向，以便它们可以与各种靶标相互作用。
Troponin C (TnC) is an 18 kDa (162-residue) thin-filament calcium-binding protein responsible for triggering muscle contraction upon the release of calcium from the sarcoplasmic reticulum. The structure of TnC with two calcium ions bound has previously been solved by X-ray methods. Shown here is the solution structure of TnC which has been solved using 3D and 4D heteronuclear nuclear magnetic resonance (NMR) spectroscopic techniques. The 1H, 13C, and 15N backbone chemical shifts have already been published [Slupsky, C. M., Reinach, F. C., Smillie, L. B., & Sykes, B. D. (1995) Protein Sci. 4, 1279-1290]. Presented herein are the 1H, 13C, and 15N side-chain chemical shifts which are 80% complete. The structure of calcium-saturated TnC was determined on the basis of 2106 NOE-derived distance restraints, 121 phi dihedral angle restraints, and 76 psi dihedral angle restraints. The appearance of calcium-saturated TnC reveals a dumbbell-shaped molecule with two globular domains connected by a linker. The structures of the N-terminal and C-terminal domains are highly converged [backbone atomic root mean square deviations (rmsd) about the mean atomic coordinate position for residues 10-80 and 98-155 are 0.66 +/- 0.17 and 0.69 +/- 0.18 A, respectively]; however, the orientation of one domain with respect to the other is not well-defined, and thus each domain appears to be structurally independent. Comparison of the calcium-saturated form of TnC determined herein with the half-saturated form determined by X-ray methods reveals two major differences. First, there is a major structural change which occurs in the N-terminal domain resulting in the opening of a hydrophobic pocket presumably to present itself to its target protein troponin I. This structural change appears to involve only helices B and C which move away from helices N/A/D by the alteration of the backbone phi, psi angles of glutamic acid 41 from irregular in the crystal structure (-97 degrees, -7 degrees) to helical in the NMR calcium-saturated structure (-60 degrees, -34 degrees). The other difference between the two structures is the presence of a flexible linker between the two domains in the NMR structure. This flexible linker allows the two domains of TnC to adopt any orientation with respect to one another such that they can interact with a variety of targets.