Brath Ulrika, Akke Mikael, Yang Daiwen, Kay Lewis E, Mulder Frans A A
Department of Biophysical Chemistry, Lund University, Box 124, SE-221 00 Lund, Sweden.
J Am Chem Soc. 2006 May 3;128(17):5718-27. doi: 10.1021/ja0570279.
Transverse relaxation dispersion NMR spectroscopy can provide atom-specific information about time scales, populations, and the extent of structural reorganization in proteins under equilibrium conditions. A method is described that uses side-chain methyl groups as local reporters for conformational transitions taking place in the microsecond regime. The experiment measures carbon nuclear spin relaxation rates in the presence of continuous wave off-resonance irradiation, in proteins uniformly enriched with 13C, and partially randomly labeled with 2H. The method was applied to human FK-506 binding protein (FKBP12), which uses a common surface for binding substrates in its dual role as both an immunophilin and folding assistant. Conformational dynamics on a time scale of approximately 130 micros were detected for methyl groups located in the substrate binding pocket, demonstrating its plasticity in the absence of substrate. The spatial arrangement of affected side-chain atoms suggests that substrate recognition involves the rapid relative movement of the subdomain comprising residues Ala81-Thr96 and that the observed dynamics play an important role in facilitating the interaction of this protein with its many partners, including calcineurin.
横向弛豫分散核磁共振波谱能够在平衡条件下提供有关蛋白质中时间尺度、种群以及结构重组程度的原子特异性信息。本文描述了一种方法，该方法使用侧链甲基作为微秒级构象转变的局部报告基团。该实验在连续波偏共振照射下，对均匀富集13C且部分随机标记2H的蛋白质中的碳核自旋弛豫率进行测量。该方法应用于人类FK-506结合蛋白（FKBP12），它作为亲免素和折叠辅助因子，在其双重作用中使用共同表面结合底物。对于位于底物结合口袋中的甲基，检测到了约130微秒时间尺度上的构象动力学，证明了其在无底物时的可塑性。受影响的侧链原子的空间排列表明，底物识别涉及包含Ala81 - Thr96残基的亚结构域的快速相对移动，并且观察到的动力学在促进该蛋白与其许多伙伴（包括钙调神经磷酸酶）的相互作用中起着重要作用。
