Bernadó Pau, Blackledge Martin
Institut de Biologie Structurale Jean-Pierre Ebel, UJF-CNRS-CEA, 41 rue Jules Horowitz, 38027 Grenoble Cedex, France.
J Am Chem Soc. 2004 Apr 21;126(15):4907-20. doi: 10.1021/ja036977w.
The effect of small amplitude anisotropic peptide plane motion on residual dipolar couplings (RDC) measured in proteins has been investigated. RDC averaging effects in the presence of GAF (Gaussian axial fluctuation) motions are found to vary strongly depending on the peptide plane orientation. Even low amplitude dynamics can significantly affect derived alignment tensor parameters if this motion is not taken into account. An analytical description of averaged N-(N)H RDCs is introduced that includes basic GAF-like motion. The averaging depends on the orientation of the peptide plane (alpha, beta, gamma) in the alignment frame and on the motional amplitude (sigma). This expression is used to investigate the presence of anisotropic reorientational dynamics in proteins by incorporating sigma as an additional parameter into the alignment tensor analysis. Average GAF amplitudes (sigma(av)) are determined for secondary structural elements from single experimental N-(N)H RDC data sets from five different proteins, in combination with high-resolution structural models. This yields statistically significant improvement over the static description, and detects sigma(av) values ranging from 14.4 to 17.0 degrees for the different proteins. A higher value of sigma(av) = 20 degrees from loop regions was found using two independent sets of N-(N)H RDC in the protein lysozyme, for which a very high-resolution structure is available. Comparison of fitting behavior over 13 structures from lysozyme of crystal diffraction resolution ranging from 0.9 to 2.1A indicates a small spread of motional amplitudes, demonstrating that the method is robust up to this level of resolution. A combined definition of (alpha)C-C' and N-(N)H RDC under the influence of GAF motions allows simultaneous fitting of both RDC. Application to three proteins leads to similar sigma(av) values and a more significant improvement with respect to the static model. Using the GAF model to describe conformationally averaged RDC is important for two reasons: a more accurate definition of the alignment tensor magnitude can be derived, and the method can be used to detect average small amplitude motions in protein backbones from readily accessible data, on time scales not easily sampled by other NMR techniques.
研究了小幅度各向异性肽平面运动对蛋白质中测量的剩余偶极耦合（RDC）的影响。发现在存在高斯轴向涨落（GAF）运动的情况下，RDC平均效应强烈依赖于肽平面的取向。如果不考虑这种运动，即使是低幅度动力学也会显著影响推导的取向张量参数。引入了对平均N - （N）H RDC的解析描述，其中包括基本的类GAF运动。平均效应取决于肽平面在取向框架中的取向（α、β、γ）以及运动幅度（σ）。该表达式用于通过将σ作为额外参数纳入取向张量分析来研究蛋白质中各向异性重排动力学的存在。结合高分辨率结构模型，从五个不同蛋白质的单个实验N - （N）H RDC数据集中确定二级结构元件的平均GAF幅度（σ(av)）。这相对于静态描述在统计上有显著改进，并检测到不同蛋白质的σ(av)值范围为14.4至17.0度。在溶菌酶中使用两组独立的N - （N）H RDC发现环区域的σ(av)值更高，为20度，其具有非常高分辨率的结构。对晶体衍射分辨率范围为0.9至2.1 Å的溶菌酶的13种结构的拟合行为进行比较，表明运动幅度的差异较小，证明该方法在该分辨率水平下是稳健的。在GAF运动影响下（α）C - C'和N - （N）H RDC的组合定义允许同时拟合这两种RDC。应用于三种蛋白质导致类似的σ(av)值，并且相对于静态模型有更显著的改进。使用GAF模型描述构象平均RDC很重要，原因有两个：可以得出取向张量大小的更准确定义，并且该方法可用于从易于获取的数据中检测蛋白质主链中的平均小幅度运动，这些时间尺度是其他NMR技术不易采样的。
