Glycerol-enhanced detection of a preferential structure latent in unstructured 1SS-variants of lysozyme
Glycerol-enhanced detection of a preferential structure latent in unstructured 1SS-variants of lysozyme
复制标题
DOI：
10.1002/bip.22037
复制
发表时间：
2012-07-01
期刊：
影响因子：
2.9
通讯作者：
Segawa, Shin-ichi
中科院分区：
文献类型：
--
作者：
Noda, Yasuo;Narama, Kuniaki;Segawa, Shin-ichi
从近紫外圆二色谱和~1H-15N-HSQC谱判断，四种溶菌酶的1SS-变异体在水中几乎是无结构的。在这种无序状态下可能存在一些择优结构，但这种构象中的分子数量肯定太小，无法用光谱方法检测到。事实上，我们之前的研究表明，添加30%的甘油可以诱导溶菌酶的非结构2SS变体形成类似天然结构的结构。为了将这种方法扩展到更多的无序蛋白质，我们试图通过甘油增强检测来检测非结构1SS变异体中潜在的一些优先结构。在含有Cys6-Cys127单二硫键的四个1SS变异体中，只有一种分子物种1SS[6-127]在50%甘油存在下具有择优结构。通过近紫外圆二次测量和氢/氢交换法结合核磁共振光谱对其进行了检测。1SS[6-127]中甘油诱导的结构不仅定位在Cys6-Cys127附近，而且主要保护区分布在A-螺旋、B-螺旋、C-螺旋以及Ile55和Leu56之间。它与甘油诱导的2SS[6-127，-80]中含有两个二硫键Cys6-Cys127和Cys64-Cys80的结构相似，尽管前者没有后者那么刚性。A-螺旋(残基415)的作用被认为是Cys6-Cys127在a-结构域诱导三级结构的良好潜力的来源。(C)2012年威利期刊公司生物聚合物97：539549,2012年。
Four species of 1SS-varinats of lysozyme were almost unstructured in water, judged from their near-UV CD and 1H-15N-HSQC spectra. Some preferential structure might exist in such a disordered state, but the population of molecules in such a conformation must have been too small to be detected by spectroscopic methods. Indeed, our previous study showed that the addition of 30% glycerol induced the unstructured 2SS-variant of lysozyme to form a native-like structure. To extend this method to more disordered proteins, we attempted to detect some preferential structure latent in unstructured 1SS-variants by the glycerol-enhanced detection. Only in one molecular species of the four 1SS-variants, 1SS[6-127] containing a single disulfide bridge of Cys6-Cys127, a preferential structure was found in the presence of 50% glycerol. It was detected by near-UV CD measurements and the H/D exchange method combined with the NMR spectroscopy. The glycerol-induced structure in 1SS[6-127] was not localized only in the vicinity of Cys6-Cys127, and largely protected regions distributed themselves among A-, B-, and C-helices and Ile55 and Leu56. It was similar to the glycerol-induced structure in 2SS[6-127, 64-80] containing two disulfide bridges of Cys6-Cys127 and Cys64-Cys80, although the former was less rigid than the latter. The role of A-helix (residues 415) is proposed as an origin of excellent potential of Cys6-Cys127 for inducing a tertiary structure in the a-domain. (c) 2012 Wiley Periodicals, Inc. Biopolymers 97:539549, 2012.