An improved 15N relaxation dispersion experiment for the measurement of millisecond time-scale dynamics in proteins
An improved 15N relaxation dispersion experiment for the measurement of millisecond time-scale dynamics in proteins
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DOI：
10.1021/jp074793o
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发表时间：
2008-05-15
影响因子：
3.3
通讯作者：
Kay, Lewis E.
中科院分区：
文献类型：
--
作者：
Hansen, D. Flemming;Vallurupalli, Pramodh;Kay, Lewis E.
提出了一种新的（15）N恒时弛豫色散脉冲方案，用于蛋白质毫秒时间尺度交换动力学的定量研究。该实验与先前开发的序列的不同之处在于，它包括（1）在（15）N Carr-Purcell-Meiboom-Gill（CPMG）脉冲串期间的H连续波去耦，其显著改善了（15）N磁化的弛豫特性，从而导致实验中的灵敏度增益。此外，示出了在由（15）N个180（y）度脉冲组成的CPMG脉冲串的中心包括额外的（15）N个180度重聚焦脉冲（相位循环+/- x），提供了对超出正常CPMG方案的脉冲缺陷的补偿。相对于现有的弛豫补偿的恒定时间弛豫色散脉冲方案，可以采用仅为一半大的v（CPMG）值，从而提供对慢时间尺度交换过程的增加的灵敏度。该方法的鲁棒性与涉及一对蛋白质的应用程序进行说明：SH 3域，不显示毫秒时间尺度的交换和FF域具有显着的化学交换贡献。
A new (15)N constant-time relaxation dispersion pulse scheme for the quantification of millisecond time-scale exchange dynamics in proteins is presented. The experiment differs from previously developed sequences in that it includes (1)H continuous-wave decoupling during the (15)N Carr-Purcell-Meiboom-Gill (CPMG) pulse train that significantly improves the relaxation properties of (15)N magnetization, leading to sensitivity gains in experiments. Moreover, it is shown that inclusion of an additional (15)N 180 degrees refocusing pulse (phase cycled +/- x) in the center of the CPMG pulse train, consisting of (15)N 180(y)degrees pulses, provides compensation for pulse imperfections beyond the normal CPMG scheme. Relative to existing relaxation-compensated constant-time relaxation dispersion pulse schemes, v(CPMG) Values that are only half as large can be employed, offering increased sensitivity to slow time-scale exchange processes. The robustness of the methodology is illustrated with applications involving a pair of proteins: an SH3 domain that does not show millisecond time-scale exchange and an FF domain with significant chemical exchange contributions.