Monitoring aromatic picosecond to nanosecond dynamics in proteins via 13C relaxation:: Expanding perturbation mapping of the rigidifying core mutation, V54A, in Eglin C
Monitoring aromatic picosecond to nanosecond dynamics in proteins via 13C relaxation:: Expanding perturbation mapping of the rigidifying core mutation, V54A, in Eglin C
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DOI：
10.1021/bi702330t
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发表时间：
2008-04-29
期刊：
影响因子：
2.9
通讯作者：
Lee, Andrew L.
中科院分区：
文献类型：
--
作者：
Boyer, Joshua A.;Lee, Andrew L.
长程效应，如变构，已经在蛋白质中进化为通过远端位点之间的通信来调节功能的手段。基于NMR的微扰映射方法被用来更全面地探测的核心突变V54 A的动态响应的蛋白质eglin c通过监测皮秒到纳秒的芳香族侧链动力学和H/D交换稳定性的变化。以前对这种突变体的侧链动力学研究仅限于含甲基的残基，其中大部分被发现在皮秒至纳秒的时间尺度上以连续的“网络”形式硬化。在此，通过将正则弛豫实验应用于新开发的碳标记方案来获得来自13个芳族侧链的高精度C-13弛豫数据[Teilum等人（2006）J. Am. 128，2506-2507]。无模型参数的拟合产生了S-2变异性，这是中间相对于骨干和甲基轴承侧链的变异性和tau（e）值，约为INS。列入的芳香族动态响应的结果，在一个扩大的网络动态耦合残基，与一些芳香族化合物显示出增加的灵活性，这部分抵消了甲基侧链的硬化。使用酰胺氢交换，在较慢的时间尺度上的动态传播探测响应于V54 A扰动。令人惊讶的是，在离突变位点10-12埃的区域内观察到了稳定（交换减慢），尽管整体不稳定为1.5 kcal.mol（-1）。此外，这种不太可能的口袋稳定的残留物共定位与芳香族的灵活性增加更快的时间尺度。因为匡威也是真的（不稳定的残留物与硬化的快速时间尺度上的共定位），一个似是而非的熵驱动的机制进行了讨论，有关共定位的对立动态趋势在截然不同的时间尺度上。
Long-range effects, such as allostery, have evolved in proteins as a means of regulating function via communication between distal sites. An NMR-based perturbation mapping approach was used to more completely probe the dynamic response of the core mutation V54A in the protein eglin c by monitoring changes in picosecond to nanosecond aromatic side-chain dynamics and H/D exchange stabilities. Previous side-chain dynamics studies on this mutant were limited to methyl-bearing residues, most of which were found to rigidify on the picosecond to nanosecond time scale in the form of a contiguous "network". Here, high precision C-13 relaxation data from 13 aromatic side chains were acquired by applying canonical relaxation experiments to a newly developed carbon labeling scheme [Teilum et al. (2006) J. Am. Chem. Soc. 128, 2506-2507]. The fitting of model-free parameters yielded S-2 variability which is intermediate with respect to backbone and methyl-bearing side-chain variability and tau(e) values that are approximately Ins. Inclusion of the aromatic dynamic response results in an expanded network of dynamically coupled residues, with some aromatics showing increases in flexibility, which partially offsets the rigidification in methyl side chains. Using amide hydrogen exchange, dynamic propagation on a slower time scale was probed in response to the V54A perturbation. Surprisingly, regional stabilization (slowed exchange) 10-12 angstrom from the site of mutation was observed despite a global destabilization of 1.5 kcal.mol(-1). Furthermore, this unlikely pocket of stabilized residues colocalizes with increases in aromatic flexibility on the faster time scale. Because the converse is also true (destabilized residues colocalize with rigidification on the fast time scale), a plausible entropy-driven mechanism is discussed for relating colocalization of opposing dynamic trends on vastly different time scales.