Estimation of inter-residue distances in spin labeled proteins at physiological temperatures: Experimental strategies and practical limitations
Estimation of inter-residue distances in spin labeled proteins at physiological temperatures: Experimental strategies and practical limitations
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DOI：
10.1021/bi011544w
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发表时间：
2001-12-25
期刊：
影响因子：
2.9
通讯作者：
Hubbell, WL
中科院分区：
文献类型：
--
作者：
Altenbach, C;Oh, KJ;Hubbell, WL
研究了T4溶菌酶中沿α -螺旋约一至四匝的暴露于溶剂中的氮侧链对之间的磁偶极相互作用。分析了冻结溶液（刚性晶格条件）和室温下溶剂粘度的相互作用。在室温下，使用了一种具有阻碍内部运动的新型侧链，以及更常用的氮氧化物侧链。结果表明，在刚性晶格条件下开发的方法可以用于分析氮氧化物之间的偶极相互作用，即使在存在单个自旋运动的情况下，只要自旋间矢量的旋转相关时间足够长。观察到的各种自旋对的距离分布与晶体结构中观察到的氮氧化物侧链的旋美平衡相一致。这种距离分布的存在对蛋白质结构和结构变化方面氮氧化物间距离的解释产生了重要的限制。
Magnetic dipolar interactions between pairs of solvent-exposed nitroxide side chains separated by approximately one to four turns along an alpha-helix in T4 lysozyme are investigated. The interactions are analyzed both in frozen solution (rigid lattice conditions) and at room temperature as a function of solvent viscosity. At room temperature, a novel side chain with hindered internal motion is used, along with a more commonly employed nitroxide side chain. The results suggest that methods developed for rigid lattice conditions can be used to analyze dipolar interactions between nitroxides even in the presence of motion of the individual spins, provided the rotational correlation time of the interspin vector is sufficiently long. The distribution of distances observed for the various spin pairs is consistent with rotameric equilibria in the nitroxide side chain, as observed in crystal structures. The existence of such distance distributions places important constraints on the interpretation of internitroxide distances in terms of protein structure and structural changes.