Interaction forces between F-Actin and titin PEVK domain measured with optical tweezers
Interaction forces between F-Actin and titin PEVK domain measured with optical tweezers
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DOI：
10.1529/biophysj.107.106153
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发表时间：
2007-09-01
影响因子：
3.4
通讯作者：
Kellermayer, Miklos S. Z.
中科院分区：
文献类型：
--
作者：
Bianco, Pasquale;Nagy, Attila;Kellermayer, Miklos S. Z.
Titin是一种巨大的蛋白质，决定横纹肌的弹性，并被认为在许多调节过程中发挥重要作用。先前的研究表明，titin的PEVK结构域与F- actin相互作用，从而产生未知大小的粘稠力，可能调节肌肉收缩。在这里，我们用光学镊子测量了将F-肌动蛋白从富含polyye或PPAK基元的重组PEVK片段的单个分子中分离出来所需的力。在拉伸速率为250 nm/s时，断裂力呈宽的非正态分布，两种碎片的峰值均接近8 pN。动态力谱实验表明，即使在较低的力作用下，自发脱轨率也较低。与PPAK观察到的单相响应相比，polyye的破裂力加载速率依赖是双相的。对发生断裂的分子长度的分析表明，PEVK片段的轮廓上有许多肌动蛋白结合区域，这表明PEVK结构域是一个混杂的肌动蛋白结合伙伴。PEVK-actin相互作用的复杂性指出了一种适应性粘弹性机制，该机制在松弛状态下保护肌肉结构的完整性，并在收缩时调节触变行为。
Titin is a giant protein that determines the elasticity of striated muscle and is thought to play important roles in numerous regulatory processes. Previous studies have shown that titin's PEVK domain interacts with F- actin, thereby creating viscous forces of unknown magnitude that may modulate muscle contraction. Here we measured, with optical tweezers, the forces necessary to dissociate F- actin from individual molecules of recombinant PEVK fragments rich either in polyE or PPAK motifs. Rupture forces at a stretch rate of 250 nm/s displayed a wide, nonnormal distribution with a peak at similar to 8 pN in the case of both fragments. Dynamic force spectroscopy experiments revealed low spontaneous off-rates that were increased even by low forces. The loading-rate dependence of rupture force was biphasic for polyE in contrast with the monophasic response observed for PPAK. Analysis of the molecular lengths at which rupture occurred indicated that there are numerous actin-binding regions along the PEVKfragments' contour, suggesting that the PEVK domain is a promiscuous actin-binding partner. The complexity of PEVK-actin interaction points to an adaptable viscoelastic mechanism that safeguards sarcomeric structural integrity in the relaxed state and modulates thixotropic behavior during contraction.