Steensma E, van Mierlo C P
Department of Biomolecular Sciences - Laboratory of Biochemistry, Wageningen Agricultural University, Dreijenlaan 3, Wageningen, NL-6703 HA, The Netherlands.
J Mol Biol. 1998 Sep 25;282(3):653-66. doi: 10.1006/jmbi.1998.2045.
The structural characteristics of Azotobacter vinelandii apoflavodoxin II have been determined using multidimensional NMR spectroscopy. Apoflavodoxin has a stable, well-ordered core but its flavin binding region is flexible. The local stability of apoflavodoxin was probed using hydrogen/deuterium exchange measurements. The existence of an apoflavodoxin equilibrium folding intermediate is inferred from the non-coincidence of CD and fluorescence unfolding curves obtained for the guanidinium hydrochloride induced unfolding of apoflavodoxin. We suggest that the structured part of the putative intermediate is composed of the elements of secondary structure which have the slowest exchanging amide protons in the native protein. These elements are strands beta1, beta3, beta4 and beta5a and helices alpha4 and alpha5. We propose that it is a general feature of flavodoxins that the stable nucleus resides in the C-terminal part of these proteins. The results on flavodoxin are compared with those on two sequentially unrelated proteins sharing the flavodoxin-like fold: Che Y and cutinase. It is shown that the stable nucleus is found in different parts of the flavodoxin-like topology.
运用多维核磁共振波谱法测定了棕色固氮菌脱辅基黄素氧还蛋白II的结构特征。脱辅基黄素氧还蛋白具有一个稳定、有序的核心，但它的黄素结合区域是灵活可变的。通过氢/氘交换测量来探究脱辅基黄素氧还蛋白的局部稳定性。从盐酸胍诱导脱辅基黄素氧还蛋白展开时获得的圆二色光谱（CD）和荧光展开曲线不重合的情况推断，脱辅基黄素氧还蛋白存在平衡折叠中间体。我们认为，假定中间体的结构化部分由天然蛋白质中酰胺质子交换最慢的二级结构元件组成。这些元件包括β1、β3、β4和β5a链以及α4和α5螺旋。我们提出，黄素氧还蛋白的一个普遍特征是稳定的核心位于这些蛋白质的C末端部分。将黄素氧还蛋白的结果与另外两种具有黄素氧还蛋白样折叠但序列不相关的蛋白质——趋化因子Y和角质酶的结果进行了比较。结果表明，在黄素氧还蛋白样拓扑结构的不同部分发现了稳定的核心。
