Resolution of three structural states of spin-labeled myosin in contracting muscle.
Resolution of three structural states of spin-labeled myosin in contracting muscle.
复制标题
解析收缩肌肉中自旋标记肌球蛋白的三种结构状态。
DOI：
10.1016/s0006-3495(95)79888-5
复制
发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Thomas,DD
中科院分区：
文献类型：
--
作者：
Ostap,EM;Barnett,VA;Thomas,DD
我们已经使用电子顺磁共振（EPR）光谱检测ATP和钙诱导的结构的变化，自旋标记的肌球蛋白头在甘油兔腰大肌肌纤维在关键的生理状态。探针是一个氮氧碘乙酰胺衍生物选择性地连接到肌球蛋白SH 1（Cys 707），传统的EPR光谱已被证明解决几个构象状态的肌球蛋白ATP酶循环，纳秒旋转运动的基础上，在蛋白质。光谱采集在僵硬和在稳态阶段的放松和等长收缩。对应于特定的构象状态和生化中间体的光谱成分被检测到，并通过引用被困的动力学中间体的EPR谱分配。在没有ATP的情况下，所有的肌球蛋白头刚性连接到细丝，只有一个单一的构象被检测到，其中没有亚微秒的探针运动。在放松，EPR谱解决了两个构象的肌球蛋白头是不同的僵硬。这些结构状态实际上与以前观察到的分离肌球蛋白的结构状态相同，并被分配到M*.ATP和M**.ADP.Pi状态的群体。在等长收缩期间，EPR谱解析了在松弛中观察到的相同的两种构象，加上在僵硬中观察到的定向肌动蛋白结合构象中的一小部分（20-30%）头部。这种僵硬样成分是一种钙依赖性肌动蛋白结合状态，可能代表产生力的跨桥。由于自旋标签位于附近的核苷酸结合口袋中的一个区域提出是关键的大规模的力产生肌球蛋白的结构变化，我们建议，所观察到的光谱变化直接表明在收缩肌肉的分子马达中的能量转导的关键步骤。
We have used electron paramagnetic resonance (EPR) spectroscopy to detect ATP- and calcium-induced changes in the structure of spin-labeled myosin heads in glycerinated rabbit psoas muscle fibers in key physiological states. The probe was a nitroxide iodoacetamide derivative attached selectively to myosin SH1 (Cys 707), the conventional EPR spectra of which have been shown to resolve several conformational states of the myosin ATPase cycle, on the basis of nanosecond rotational motion within the protein. Spectra were acquired in rigor and during the steady-state phases of relaxation and isometric contraction. Spectral components corresponding to specific conformational states and biochemical intermediates were detected and assigned by reference to EPR spectra of trapped kinetic intermediates. In the absence of ATP, all of the myosin heads were rigidly attached to the thin filament, and only a single conformation was detected, in which there was no sub-microsecond probe motion. In relaxation, the EPR spectrum resolved two conformations of the myosin head that are distinct from rigor. These structural states were virtually identical to those observed previously for isolated myosin and were assigned to the populations of the M*.ATP and M**.ADP.Pi states. During isometric contraction, the EPR spectrum resolves the same two conformations observed in relaxation, plus a small fraction (20–30%) of heads in the oriented actin-bound conformation that is observed in rigor. This rigor-like component is a calcium-dependent, actin-bound state that may represent force-generating cross-bridges. As the spin label is located near the nucleotide-binding pocket in a region proposed to be pivotal for large-scale force-generating structural changes in myosin, we propose that the observed spectroscopic changes indicate directly the key steps in energy transduction in the molecular motor of contracting muscle.
登录
查看更多内容
影响因子：
3.4
作者：
David D. Thomas;Sampath Ramachandran;O. Roopnarine;David W. Hayden;E. Ostap
通讯作者：
David D. Thomas;Sampath Ramachandran;O. Roopnarine;David W. Hayden;E. Ostap
活跃肌原纤维中肌动蛋白结合的肌球蛋白头的旋转动力学。
影响因子：
2.9
作者：
Berger,CL;Thomas,DD
通讯作者：
Thomas,DD
N-乙基马来酰亚胺修饰肌球蛋白的酶学特性
DOI：
--
发表时间：
1964
期刊：
影响因子：
--
作者：
T. Sekine;W. W. Kielley
通讯作者：
W. W. Kielley
使用单纯形算法对高分辨率 EPR 谱进行多参数拟合的通用方法☆
DOI：
--
发表时间：
1990
期刊：
影响因子：
--
作者：
P. Fajer;R. Bennett;C. Polnaszek;E. A. Fajer;David D. Thomas
通讯作者：
David D. Thomas
肌球蛋白亚片段的乙烯腺苷二磷酸结合位点的表征 1. 通过能量转移检测到的核苷酸两种状态之间平衡的能量。S1 和钒酸盐诱导的整体构象变化。
影响因子：
2.9
作者：
Aguirre,R;Lin,SH;Gonsoulin,F;Wang,CK;Cheung,HC
通讯作者：
Cheung,HC