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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• 作者：Jennifer B. Hall and David Fushman
• 刊名：Journal of the American Chemical Society
• 出版年：2006
• 出版时间：June 21, 2006
• 年：2006
• 卷：128
• 期：24
• 页码：7855 - 7870
• 全文大小：333K
• 年卷期：v.128,no.24(June 21, 2006)
• ISSN：1520-5126

文摘

We applied a combination of ¹⁵N relaxation and CSA/dipolar cross-correlation measurements atfive magnetic fields (9.4, 11.7, 14.1, 16.4, and 18.8 T) to determine the ¹⁵N chemical shielding tensors forbackbone amides in protein G in solution. The data were analyzed using various model-independentapproaches and those based on Lipari-Szabo approximation, all of them yielding similar results. The resultsindicate a range of site-specific values of the anisotropy (CSA) and orientation of the ¹⁵N chemical shieldingtensor, 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 Å NH bond length) and the site-to-site CSA variability is ±17.6 to ±21.4ppm, depending on the method used. This CSA variability is significantly larger than derived previously forribonuclease 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 ¹⁵N relaxationstudies of backbone dynamics in proteins involves an a priori assumption of a uniform ¹⁵N CSA. We showthat this assumption leads to a significant discrepancy between the order parameters obtained at differentfields. Using the site-specific CSAs obtained from our study removes this discrepancy and allowssimultaneous fit of relaxation data at all five fields to Lipari-Szabo spectral densities. These findingsemphasize the necessity of taking into account the variability of ¹⁵N CSA for accurate analysis of proteindynamics from ¹⁵N relaxation measurements.