An unlocking/relocking barrier in conformational fluctuations of villin headpiece subdomain
An unlocking/relocking barrier in conformational fluctuations of villin headpiece subdomain
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DOI：
10.1073/pnas.0910001107
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发表时间：
2010-03-16
影响因子：
11.1
通讯作者：
Kiefhaber, Thomas
中科院分区：
文献类型：
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作者：
Reiner, Andreas;Henklein, Peter;Kiefhaber, Thomas
利用三重态-三重态-能量转移监测平衡态的构象波动，发现绒毛蛋白头片段亚结构域（HP 35）存在一个可逆的结构解锁反应，其中紧密堆积的货车德瓦尔斯相互作用协同破坏。解锁与不利的焓变（Δ H（0）= 35 +/-4 kJ/mol）相关，其在自由能方面几乎被熵变（Δ S（0）= 112 +/-20 J）补偿。mol（-1）。K（-1））。解锁反应在5 ℃下的时间常数约为1 μ s，并且是可逆限制的，活化能为32 +/- 1 kJ/mol，大的Arrhenius指前因子为A = 7.5 × 10（11）s（-1）。在解锁状态下，快速的局部构象波动与170 ns的时间常数和17 +/- 1 kJ/mol的低活化势垒导致C-末端螺旋的展开和其从核心的解对接。在慢得多的时间尺度上，全局展开从解锁状态发生。这些结果表明，天然蛋白质结构被锁定到低幅度运动的构象。大尺度运动和全球展开需要一个初始的结构解锁步骤，导致一个干燥的熔融球的属性的状态。实验还产生了关于未折叠的HP 35中不同位置之间的环形成的动力学的信息。与非结构化模型肽的动力学的结果比较表明，在C-末端区域的点在残留的，非随机结构的局部循环形成的动力学略有减速。相反，长程循环形成的动力学不受残余结构的影响，这与作为HP 35超快折叠的分子起源的未折叠状态属性相矛盾。
A reversible structural unlocking reaction, in which the close-packed van der Waals interactions break cooperatively, has been found for the villin headpiece subdomain (HP35) using triplet-triplet-energy transfer to monitor conformational fluctuations from equilibrium. Unlocking is associated with an unfavorable enthalpy change (Delta H(0) = 35 +/- 4 kJ/mol) which is nearly compensated in free energy by the entropy change (Delta S(0) = 112 +/- 20 J . mol(-1) . K(-1)). The unlocking reaction has a time constant of about 1 mu s at 5 degrees C and is enthalpy-limited with an activation energy of 32 +/- 1 kJ/mol and a large Arrhenius preexponential factor of A = 7.5 x 10(11) s(-1). In the unlocked state a fast local conformational fluctuation with a time constant of 170 ns and a low activation barrier of 17 +/- 1 kJ/mol leads to unfolding of the C-terminal helix and to its undocking from the core. On a much slower time scale, global unfolding occurs from the unlocked state. These results suggest that native protein structures are locked into conformations with low amplitude motions. Large scale motions and global unfolding require an initial structural unlocking step leading to a state with properties of a dry molten globule. The experiments additionally yielded information on the dynamics of loop formation between different positions in unfolded HP35. Comparison of the results with dynamics in unstructured model peptides indicates slightly decelerated kinetics of local loop formation in the C-terminal region which points at residual, nonrandom structure. Dynamics of long-range loop formation, in contrast, are not influenced by residual structure, which argues against unfolded state properties as molecular origin for ultrafast folding of HP35.