Differential Responses of the Backbone and Side-Chain Conformational Dynamics in FKBP12 upon Binding the Transition-State Analog FK506: Implications for Transition-State Stabilization and Target Protein Recognition
Differential Responses of the Backbone and Side-Chain Conformational Dynamics in FKBP12 upon Binding the Transition-State Analog FK506: Implications for Transition-State Stabilization and Target Protein Recognition
复制标题
DOI：
10.1016/j.jmb.2009.01.047
复制
发表时间：
2009-03-20
影响因子：
5.6
通讯作者：
Akke, Mikael
中科院分区：
文献类型：
--
作者：
Brath, Ulrika;Akke, Mikael
FKBP12作为肽基脯氨酰顺反异构酶和作为几种细胞信号传导途径的调节剂发挥双重作用。大环内酯FK506是催化反应的过渡态类似物，并从其天然靶蛋白中取代FKBP 12。我们比较了主链和主链的构象交换动力学。FKBP 12的甲基侧链在自由和FK506结合状态下使用NMR弛豫分散实验。我们的研究结果表明，自由的酶之间的基态和激发态，类似于配体结合态或米氏复合物的交流。在FK506结合的FKBP12中，骨架被限制为单一构象，而许多甲基基团普遍存在构象交换。在过渡态模拟束缚态的残余侧链动力学表明，过渡态系综涉及多种构象，这一发现挑战了长期存在的概念，在过渡态复合物的构象限制。此外，在束缚状态下观察到替代构象之间的交换，用于包括远离活性位点的位置的甲基基团的扩展网络。已知这些位置中的几个对于与细胞靶蛋白（包括钙调磷酸酶和ryanodine受体）的相互作用是重要的，这表明构象异质性可能在FKBP 12与不同靶标的混杂结合中起作用。(C)2009爱思唯尔有限公司版权所有。
FKBP12 serves a dual role as a peptidyl-prolyl cis-trans isomerase and as a modulator of several cell signaling pathways. The macrolide FK506 is a transition-state analog of the catalyzed reaction and displaces FKBP12 from its natural target proteins. We compared the conformational exchange dynamics of the backbone and. methyl-bearing side chains of FKBP12 in the free and FK506-bound states using NMR relaxation-dispersion experiments. Our results show that the free enzyme exchanges between the ground state and an excited state that resembles the ligand-bound state or Michaelis complex. In FK506-bound FKBP12, the backbone is confined to a single conformation, while conformational exchange prevails for many methyl groups. The residual side-chain dynamics in the transition-state analog-bound state suggests that the transition-state ensemble involves multiple conformations, a finding that challenges the long-standing concept of conformational restriction in the transition-state complex. Furthermore, exchange between alternative conformations is observed in the bound state for an extended network of methyl groups that includes locations remote from the active site. Several of these locations are known to be important for interactions with cellular target proteins, including calcineurin and the ryanodine receptor, suggesting that the conformational heterogeneity might play a role in the promiscuous binding of FKBP12 to different targets. (C) 2009 Elsevier Ltd All rights reserved.