Monitoring macromolecular motions on microsecond to millisecond time scales by R(1)rho-R(1) constant relaxation time NMR spectroscopy
Monitoring macromolecular motions on microsecond to millisecond time scales by R(1)rho-R(1) constant relaxation time NMR spectroscopy
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DOI：
10.1021/ja953503r
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发表时间：
1996-01-31
影响因子：
15
通讯作者：
Palmer, AG
中科院分区：
文献类型：
--
作者：
Akke, M;Palmer, AG
在微秒到毫秒（µs-ms）时间尺度上的动态过程对蛋白质的功能非常重要，包括识别、变构和催化。1,2分子内运动在µs-ms时间尺度上通过相干态的绝热减相导致核磁弛豫，并在溶液态核磁共振波谱中表现为构象交换现象。旋转框架中的核磁弛豫（即在射频（rf）场存在的情况下）构成了化学和构象交换过程的独特信息来源。本文提出了一种新的旋转框架技术，用于研究蛋白质5-8分子内和分子间的交换，克服了与现有自旋锁定和自旋回波实验相关的几个困难。首先，在一个新的恒定弛豫时间（CRT）周期内对旋转框架和实验室框架的弛豫速率常数进行平均，以简化自旋锁实验中通常遇到的非共振效应。其次，使用非共振自旋锁射频场9-11来增加旋转框架中有效磁场的大小，以便访问更快的动态过程。非共振R1F-R1 CRT核磁弛豫实验允许测定蛋白质的构象交换时间至少短至25µs。
Dynamic processes on microsecond to millisecond (µs-ms) time scales are important for the functions of proteins, including recognition, allostery, and catalysis. 1, 2 Intramolecular motions on µs-ms time scales contribute to nuclear magnetic relaxation through adiabatic dephasing of coherent states and are exhibited as conformational exchange phenomena in solution-state NMR spectroscopy. 3 Nuclear magnetic relaxation in the rotating frame (ie, in the presence of a radiofrequency (rf) field) constitutes a unique source of information on chemical and conformational exchange processes. 4 This communication presents a new rotating frame technique for studying intra-and intermolecular exchange in proteins5-8 that overcomes several difficulties associated with existing spin-lock and spin-echo experiments. First, rotating frame and laboratory frame relaxation rate constants are averaged during a novel constant relaxation time (CRT) period in order to simplify the off-resonance effects normally encountered in spin-lock experiments. Second, an offresonance spin-lock rf field9-11 is used to increase the magnitude of the effective magnetic field in the rotating frame in order to access faster dynamic processes. The off-resonance R1F-R1 CRT nuclear magnetic relaxation experiment allows determination of conformational exchange times at least as short as 25 µs in proteins.