Water proton spin saturation affects measured protein backbone 15N spin relaxation rates
Water proton spin saturation affects measured protein backbone 15N spin relaxation rates
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DOI：
10.1016/j.jmr.2011.09.042
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发表时间：
2011-12-01
影响因子：
2.2
通讯作者：
Tjandra, Nico
中科院分区：
文献类型：
--
作者：
Chen, Kang;Tjandra, Nico
蛋白质骨架N-15 NMR自旋弛豫速率可用于表征蛋白质动力学和结构。为了观察蛋白质核自旋共振，脉冲序列必须包括水抑制方案。有两种常用的方法，饱和或移相的水自旋与脉冲场梯度和保持它们不受干扰的翻转脉冲。在这里，不同的水抑制方法被纳入脉冲序列，以测量N-15纵向T-1和横向双标架T-1 ρ自旋弛豫。出乎意料的是，N-15 T-1弛豫时间常数随水抑制方法的选择而显著变化。对于25-kDa大肠杆菌。对于谷氨酰胺结合蛋白（GlnBP），用含有水失相梯度的脉冲序列获得的T-1值比用含有水翻转脉冲的脉冲序列获得的T-1值平均长20%。相比之下，两个T-1 ρ数据集是相关的，没有明显的偏移。当实验循环延迟加倍时，平均T-1差异减小到12%，而来自翻转测量的平均T-1值几乎不变。光谱信噪比（s/n）的分析表明，明显较慢的N-15弛豫得到的水失相实验起源于H-1（N）的恢复为每个弛豫时间点的差异。这又抵消了来自N-15弛豫衰减的信号减小。当测量的N-15弛豫时间常数与再循环延迟相当时，伪影变得明显，例如，N-15 T-1的中型至大型蛋白质。N-15弛豫率测量与水抑制计划产生合理的适合的结构。然而，饱和方案的数据导致无模型阶次参数（< S-2 >= 0.81）显着低于非饱和方案（< S-2 >= 0.88），这表明此类阶次参数之前可能被低估了。爱思唯尔公司出版
Protein backbone N-15 NMR spin relaxation rates are useful in characterizing the protein dynamics and structures. To observe the protein nuclear-spin resonances a pulse sequence has to include a water suppression scheme. There are two commonly employed methods, saturating or dephasing the water spins with pulse field gradients and keeping them unperturbed with flip-back pulses. Here different water suppression methods were incorporated into pulse sequences to measure N-15 longitudinal T-1 and transversal rotating-frame T-1 rho spin relaxation. Unexpectedly the N-15 T-1 relaxation time constants varied significantly with the choice of water suppression method. For a 25-kDa Escherichia coli. glutamine binding protein (GlnBP) the T-1 values acquired with the pulse sequence containing a water dephasing gradient are on average 20% longer than the ones obtained using a pulse sequence containing the water flip-back pulse. In contrast the two T-1 rho data sets are correlated without an apparent offset. The average T-1 difference was reduced to 12% when the experimental recycle delay was doubled, while the average T-1 values from the flip-back measurements were nearly unchanged. Analysis of spectral signal to noise ratios (s/n) showed the apparent slower N-15 relaxation obtained with the water dephasing experiment originated from the differences in H-1(N) recovery for each relaxation time point. This in turn offset signal reduction from N-15 relaxation decay. The artifact becomes noticeable when the measured N-15 relaxation time constant is comparable to recycle delay, e.g., the N-15 T-1 of medium to large proteins. The N-15 relaxation rates measured with either water suppression schemes yield reasonable fits to the structure. However, data from the saturated scheme results in significantly lower Model-Free order parameters (< S-2 > = 0.81) than the non-saturated ones (< S-2 > = 0.88), indicating such order parameters may be previously underestimated. Published by Elsevier Inc.