Variability of the 15N chemical shielding tensors in the B3 domain of protein G from 15N relaxation measurements at several fields.: Implications for backbone order parameters
Variability of the 15N chemical shielding tensors in the B3 domain of protein G from 15N relaxation measurements at several fields.: Implications for backbone order parameters
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DOI：
10.1021/ja060406x
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发表时间：
2006-06-21
影响因子：
15
通讯作者：
Fushman, David
中科院分区：
文献类型：
--
作者：
Hall, Jennifer B.;Fushman, David
我们在5个磁场（9.4、11.7、14.1、16.4和18.8 T）下应用N-15弛豫和CSA/偶极互相关测量相结合的方法来确定溶液中蛋白质G中主链酰胺的15N化学屏蔽张量。使用各种模型无关的方法和基于Lipari-Szabo近似的方法分析数据，所有这些方法都得到了类似的结果。结果表明，15N化学屏蔽张量的各向异性（CSA）和取向的位点特异性值范围与泛素中的相似（Fushman等）。j。化学。社会科学学报，1998,20 (2):447；j。化学。社会科学，1999,121,8577)。假设15N CSA值为高斯分布，则各向异性的平均值为-173.9至-177.2 ppm（对于1.02埃的氢键长度），而不同地点的CSA变异性为+/- 17.6至+/- 21.4 ppm，具体取决于所使用的方法。这种CSA变异性明显大于先前推导的核糖核酸酶H （Kroenke等）。j。化学。Soc. 1999, 121, 10119)或最近，使用“元分析”泛素（Damberg, et al.）。j。化学。Soc. 2005, 127, 1995)。蛋白质骨架动力学的N-15弛豫研究的标准解释涉及一个均匀的15N CSA的先验假设。我们表明，这一假设导致了在不同场得到的序参数之间的显著差异。使用从我们的研究中获得的特定地点的csa消除了这种差异，并允许同时拟合所有五个油田的弛豫数据与Lipari-Szabo谱密度。这些发现强调了考虑N-15 CSA的可变性的必要性，以便从N-15弛豫测量中准确分析蛋白质动力学。
We applied a combination of N-15 relaxation and CSA/dipolar cross-correlation measurements at five magnetic fields (9.4, 11.7, 14.1, 16.4, and 18.8 T) to determine the 15N chemical shielding tensors for backbone amides in protein G in solution. The data were analyzed using various model-independent approaches and those based on Lipari-Szabo approximation, all of them yielding similar results. The results indicate a range of site-specific values of the anisotropy (CSA) and orientation of the 15N chemical shielding tensor, similar to those in ubiquitin (Fushman, et al. J. Am. Chem. Soc. 1998, 120, 10947; J. Am. Chem. Soc. 1999, 121, 8577). Assuming a Gaussian distribution of the 15N CSA values, the mean anisotropy is -173.9 to -177.2 ppm (for 1.02 angstrom NH bond length) and the site-to-site CSA variability is +/- 17.6 to +/- 21.4 ppm, depending on the method used. This CSA variability is significantly larger than derived previously for ribonuclease H (Kroenke, et al. J. Am. Chem. Soc. 1999, 121, 10119) or recently, using "meta-analysis" for ubiquitin (Damberg, et al. J. Am. Chem. Soc. 2005, 127, 1995). Standard interpretation of N-15 relaxation studies of backbone dynamics in proteins involves an a priori assumption of a uniform 15N CSA. We show that this assumption leads to a significant discrepancy between the order parameters obtained at different fields. Using the site-specific CSAs obtained from our study removes this discrepancy and allows simultaneous fit of relaxation data at all five fields to Lipari-Szabo spectral densities. These findings emphasize the necessity of taking into account the variability of N-15 CSA for accurate analysis of protein dynamics from N-15 relaxation measurements.