Beckman Institute, California Institute of Technology, Pasadena, CA, 91125, USA.
J Biol Inorg Chem. 2014 Jun;19(4-5):555-63. doi: 10.1007/s00775-013-1077-2. Epub 2013 Dec 31.
We present a new approach to visualizing and quantifying the displacement of segments of Pseudomonas aeruginosa azurin in the early stages of denaturation. Our method is based on a geometrical method developed previously by the authors, and elaborated extensively for azurin. In this study, we quantify directional changes in three α-helical regions, two regions having β-strand residues, and three unstructured regions of azurin. Snapshots of these changes as the protein unfolds are displayed and described quantitatively by introducing a scaling diagnostic. In accord with molecular dynamics simulations, we show that the long α-helix in azurin (residues 54-67) is displaced from the polypeptide scaffolding and then pivots first in one direction, and then in the opposite direction as the protein continues to unfold. The two β-strand chains remain essentially intact and, except in the earliest stages, move in tandem. We show that unstructured regions 72-81 and 84-91, hinged by β-strand residues 82-83, pivot oppositely. The region comprising residues 72-91 (40 % hydrophobic and 16 % of the 128 total residues) forms an effectively stationary region that persists as the protein unfolds. This static behavior is a consequence of a dynamic balance between the competing motion of two segments, residues 72-81 and 84-91.
我们提出了一种新的方法来可视化和量化铜绿假单胞菌菌绿蛋白在变性早期阶段的片段位移。我们的方法基于作者之前开发的一种几何方法，并针对菌绿蛋白进行了广泛的阐述。在这项研究中，我们定量了菌绿蛋白中三个α-螺旋区域、两个具有β-折叠残基的区域和三个无规卷曲区域的方向变化。通过引入比例诊断，我们展示并定量描述了这些变化的快照，随着蛋白质展开，这些变化会发生。与分子动力学模拟一致，我们表明菌绿蛋白中长的α-螺旋（残基 54-67）从多肽支架中位移，然后在一个方向上枢转，然后在蛋白质继续展开时向相反方向枢转。两条β-折叠链基本保持完整，并且除了在最早的阶段外，它们一起移动。我们表明，由β-折叠残基 82-83 铰接的无规卷曲区域 72-81 和 84-91 反向枢转。由残基 72-91 组成的区域（占 40%的疏水性和总残基的 16%）形成一个有效的稳定区域，在蛋白质展开过程中保持不变。这种静态行为是两个片段（残基 72-81 和 84-91）竞争运动之间的动态平衡的结果。
