Jules Stein Eye Institute, UCLA School of Medicine, Los Angeles, California 90095, USA.
J Phys Chem B. 2011 Jan 20;115(2):397-405. doi: 10.1021/jp108871m. Epub 2010 Dec 17.
In site directed spin labeling, a nitroxide side chain is introduced at a selected site in a protein; the most commonly used is a disulfide-linked side chain designated R1. The electron paramagnetic resonance (EPR) spectra of R1, and the interspin distance between pairs of R1 residues as determined by dipolar EPR spectroscopy, encode a wealth of information on the protein structure and dynamics. However, extracting this information requires structural and dynamical models of the R1 side chain, that is, the favored rotamers, the intraresidue interactions that stabilize them, and the internal modes of motion. X-ray crystal structures of R1 in proteins have revealed a set of preferred rotamers in the crystal lattice. To identify the intraresidue interactions that stabilize particular rotamers of R1 in the absence of interactions with nearby side chains in a helix, and to evaluate models for the internal motion of the side chain, quantum mechanical calculations were performed on a relevant fragment of R1 in a 10-residue α-helix. Relative rotamer energies were determined in the gas phase, and solvation energies were estimated from a continuum solvent model that includes both electrostatic and hydrophobic contributions. The results identified preferred rotamers that are in agreement with the X-ray crystallographic studies. The rotamers are apparently stabilized by intraresidue sulfur-backbone interactions, suggesting that the preferred rotamers may be the same at all solvent-exposed helix sites.
在定向自旋标记中，将氮氧自由基侧链引入蛋白质的选定位置；最常用的是一种称为 R1 的二硫键连接的侧链。R1 的电子顺磁共振（EPR）谱以及通过偶极 EPR 光谱确定的 R1 残基对之间的自旋-自旋距离，编码了有关蛋白质结构和动力学的丰富信息。然而，提取这些信息需要 R1 侧链的结构和动力学模型，即优选的构象、稳定它们的残基内相互作用以及内部运动模式。R1 在蛋白质中的 X 射线晶体结构揭示了晶体晶格中一组优选的构象。为了确定在没有与螺旋中附近侧链相互作用的情况下稳定 R1 特定构象的残基内相互作用，并评估侧链内部运动的模型，对 10 个残基α-螺旋中 R1 的相关片段进行了量子力学计算。在气相中确定了相对构象能，并从包含静电和疏水贡献的连续溶剂模型估算了溶剂化能。结果确定了与 X 射线晶体学研究一致的优选构象。构象显然是由残基内的硫-骨架相互作用稳定的，这表明在所有暴露于溶剂的螺旋位置，优选的构象可能是相同的。
