Toward Precise Interpretation of DEER-Based Distance Distributions: Insights from Structural Characterization of V1 Spin-Labeled Side Chains
Toward Precise Interpretation of DEER-Based Distance Distributions: Insights from Structural Characterization of V1 Spin-Labeled Side Chains
复制标题
DOI：
10.1021/acs.biochem.6b00608
复制
发表时间：
2016-09-20
期刊：
影响因子：
2.9
通讯作者：
Ernst, Oliver P.
中科院分区：
文献类型：
--
作者：
Balo, Aidin R.;Feyrer, Hannes;Ernst, Oliver P.
脉冲电子顺磁共振实验可以测量蛋白质中两个自旋标记侧链之间的个体距离，范围约为 1.5-8 nm。然而，传统自旋标记侧链的灵活性导致自旋密度位点扩散，从而导致峰相对较宽的距离分布，从而使蛋白质构象状态的解释复杂化。在这里，我们分析了自旋标记的 VI 侧链，它是内部锚定的，因此灵活性较差。在α-螺旋片段上携带VI侧链的V1标记的T4溶菌酶构建体的晶体结构表明VI侧链仅采用一些离散的旋转异构体。在大多数情况下，在给定位点仅观察到一个旋转异构体，这解释了双 VI 标记蛋白质经常观察到的窄距离分布。我们利用现有数据得出了指导方针，可以对其他 V1 标记的蛋白质进行距离解释，以实现更高精度的结构建模。
Pulsed electron paramagnetic resonance experiments can measure individual distances between two spin-labeled side chains in proteins in the range of similar to 1.5-8 nm. However, the flexibility of traditional spin-labeled side chains leads to diffuse spin density loci and thus distance distributions with relatively broad peaks, thereby complicating the interpretation of protein conformational states. Here we analyzed the spin-labeled VI side chain, which is internally anchored and hence less flexible. Crystal structures of V1-labeled T4 lysozyme constructs carrying the VI side chain on a-helical segments suggest that VI side chains adopt only a few discrete rotamers. In most cases, only one rotamer is observed at a given site, explaining the frequently observed narrow distance distribution for doubly VI-labeled proteins. We used the present data to derive guidelines that may allow distance interpretation of other V1-labeled proteins for higher-precision structural modeling.