Werner J M, Knott V, Handford P A, Campbell I D, Downing A K
Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK.
J Mol Biol. 2000 Mar 3;296(4):1065-78. doi: 10.1006/jmbi.1999.3513.
Calcium binding (cb) epidermal growth factor-like (EGF) domains are found in a wide variety of extracellular proteins with diverse functions. In several proteins, including the fibrillins (1 and 2), the low-density lipoprotein receptor, the Notch receptor and related molecules, these domains are organised as multiple tandem repeats. The functional importance of calcium-binding by EGF domains has been underscored by the identification of missense mutations associated with defective calcium-binding, which have been linked to human diseases. Here, we present (15)N backbone relaxation data for a pair of cbEGF domains from fibrillin-1, the defective protein in the Marfan syndrome. The data were best fit using a symmetric top model, confirming the extended conformation of the cbEGF domain pair. Our data demonstrate that calcium plays a key role in stabilising the rigidity of the domain pair on the pico- to millisecond time-scale. Strikingly, the most dynamically stable region of the construct is centred about the domain interface. These results provide important insight into the properties of intact fibrillin-1, the consequences of Marfan syndrome causing mutations, and the ultrastructure of fibrillins and other extracellular matrix proteins.
钙结合（cb）表皮生长因子样（EGF）结构域存在于多种具有不同功能的细胞外蛋白质中。在包括原纤蛋白（1和2）、低密度脂蛋白受体、Notch受体及相关分子在内的几种蛋白质中，这些结构域以多个串联重复的形式排列。与缺陷性钙结合相关的错义突变的鉴定强调了EGF结构域钙结合的功能重要性，这些突变与人类疾病有关。在此，我们展示了来自原纤蛋白-1（马凡综合征中的缺陷蛋白）的一对cbEGF结构域的（15）N主链弛豫数据。使用对称陀螺模型对数据进行了最佳拟合，证实了cbEGF结构域对的伸展构象。我们的数据表明，钙在皮秒到毫秒时间尺度上稳定结构域对的刚性方面起着关键作用。引人注目的是，构建体中最动态稳定的区域集中在结构域界面周围。这些结果为完整原纤蛋白-1的特性、马凡综合征致病突变的后果以及原纤蛋白和其他细胞外基质蛋白的超微结构提供了重要见解。
