Vugmeyster Liliya, Ostrovsky Dmitry, Villafranca Toni, Sharp Janelle, Xu Wei, Lipton Andrew S, Hoatson Gina L, Vold Robert L
University of Colorado at Denver , Denver, Colorado 80204, United States.
University of Alaska Anchorage , Anchorage, Alaska 99508, United States.
J Phys Chem B. 2015 Nov 25;119(47):14892-904. doi: 10.1021/acs.jpcb.5b09299. Epub 2015 Nov 12.
We conducted a detailed investigation of the dynamics of two phenylalanine side chains in the hydrophobic core of the villin headpiece subdomain protein (HP36) in the hydrated powder state over the 298-80 K temperature range. Our main tools were static deuteron NMR measurements of longitudinal relaxation and line shapes supplemented with computational modeling. The temperature dependence of the relaxation times reveals the presence of two main mechanisms that can be attributed to the ring-flips, dominating at high temperatures, and small-angle fluctuations, dominating at low temperatures. The relaxation is nonexponential at all temperatures with the extent of nonexponentiality increasing from higher to lower temperatures. This behavior suggests a distribution of conformers with unique values of activation energies. The central values of the activation energies for the ring-flipping motions are among the smallest reported for aromatic residues in peptides and proteins and point to a very mobile hydrophobic core. The analysis of the widths of the distributions, in combination with the earlier results on the dynamics of flanking methyl groups (Vugmeyster et al. J. Phys. Chem. B 2013, 117, 6129-6137), suggests that the hydrophobic core undergoes slow concerted fluctuations. There is a pronounced effect of dehydration on the ring-flipping motions, which shifts the distribution toward more rigid conformers. The crossover temperature between the regions of dominance of the small-angle fluctuations and ring-flips shifts from 195 K in the hydrated protein to 278 K in the dry one. This result points to the role of solvent in softening the core and highlights aromatic residues as markers of the protein dynamical transitions.
我们对水化粉末状态下的绒毛蛋白头部亚结构域蛋白（HP36）疏水核心中两个苯丙氨酸侧链在298 - 80 K温度范围内的动力学进行了详细研究。我们的主要工具是纵向弛豫和线形的静态氘核核磁共振测量，并辅以计算建模。弛豫时间的温度依赖性揭示了存在两种主要机制，可归因于高温下占主导的环翻转和低温下占主导的小角度波动。在所有温度下弛豫都是非指数性的，非指数程度从高温到低温增加。这种行为表明存在具有独特活化能值的构象体分布。环翻转运动的活化能中心值是肽和蛋白质中芳香族残基报道的最小值之一，表明疏水核心非常灵活。对分布宽度的分析，结合早期关于侧翼甲基动力学的结果（Vugmeyster等人，《物理化学杂志B》2013年，117卷，6129 - 6137页），表明疏水核心经历缓慢协同波动。脱水对环翻转运动有显著影响，使分布向更刚性构象体偏移。小角度波动和环翻转主导区域之间的交叉温度从水化蛋白中的195 K转变为干燥蛋白中的278 K。这一结果表明溶剂在软化核心中的作用，并突出芳香族残基作为蛋白质动力学转变的标记。
