Osborne M J, Wright P E
Department of Molecular Biology and Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
J Biomol NMR. 2001 Mar;19(3):209-30. doi: 10.1023/a:1011283809984.
Model-free analysis has been extensively used to extract information on motions in proteins over a wide range of timescales from NMR relaxation data. We present a detailed analysis of the effects of rotational anisotropy on the model-free analysis of a ternary complex for dihydrofolate reductase (DHFR). Our findings show that the small degree of anisotropy exhibited by DHFR (Dparallel/Dperpendicular = 1.18) introduces erroneous motional models, mostly exchange terms, to over 50% of the NH spins analyzed when isotropic tumbling is assumed. Moreover, there is a systematic change in S2, as large as 0.08 for some residues. The significant effects of anisotropic rotational diffusion on model-free motional parameters are in marked contrast to previous studies and are accentuated by lowering of the effective correlation time using isotropic tumbling methods. This is caused by the preponderance of NH vectors aligned perpendicular to the principal diffusion tensor axis and is readily detected because of the high quality of the relaxation data. A novel procedure, COPED (COmparison of Predicted and Experimental Diffusion tensors) is presented for distinguishing genuine motions from the effects of anisotropy by comparing experimental relaxation data and data predicted from hydrodynamic analyses. The procedure shows excellent agreement with the slow motions detected from the axially symmetric model-free analysis and represents an independent procedure for determining rotational diffusion and slow motions that can confirm or refute established procedures that rely on relaxation data. Our findings show that neglect of even small degrees of rotational diffusion anisotropy can introduce significant errors in model-free analysis when the data is of high quality. These errors can hinder our understanding of the role of internal motions in protein function.
无模型分析已被广泛用于从核磁共振弛豫数据中提取蛋白质在广泛时间尺度上的运动信息。我们详细分析了旋转各向异性对二氢叶酸还原酶（DHFR）三元复合物无模型分析的影响。我们的研究结果表明，DHFR表现出的小程度各向异性（D平行/D垂直 = 1.18）在假设各向同性翻滚时，会给超过50%分析的NH自旋引入错误的运动模型，主要是交换项。此外，S2存在系统性变化，某些残基的变化高达0.08。各向异性旋转扩散对无模型运动参数的显著影响与先前的研究形成鲜明对比，并且通过使用各向同性翻滚方法降低有效相关时间而更加突出。这是由于NH矢量主要垂直于主扩散张量轴排列所致，并且由于弛豫数据的高质量而易于检测到。提出了一种新的程序，即COPED（预测和实验扩散张量比较），通过比较实验弛豫数据和流体动力学分析预测的数据来区分真实运动和各向异性的影响。该程序与从轴对称无模型分析检测到的慢运动显示出极好的一致性，并且代表了一种独立的确定旋转扩散和慢运动的程序，可用于确认或反驳依赖弛豫数据的既定程序。我们的研究结果表明，当数据质量高时，即使忽略小程度的旋转扩散各向异性也会在无模型分析中引入显著误差。这些误差可能会阻碍我们对蛋白质功能中内部运动作用的理解。
