Makabe Koki, McElheny Dan, Tereshko Valentia, Hilyard Aaron, Gawlak Grzegorz, Yan Shude, Koide Akiko, Koide Shohei
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, IL 60637, USA.
Proc Natl Acad Sci U S A. 2006 Nov 21;103(47):17753-8. doi: 10.1073/pnas.0606690103. Epub 2006 Nov 8.
Although the beta-rich self-assemblies are a major structural class for polypeptides and the focus of intense research, little is known about their atomic structures and dynamics due to their insoluble and noncrystalline nature. We developed a protein engineering strategy that captures a self-assembly segment in a water-soluble molecule. A predefined number of self-assembling peptide units are linked, and the beta-sheet ends are capped to prevent aggregation, which yields a mono-dispersed soluble protein. We tested this strategy by using Borrelia outer surface protein (OspA) whose single-layer beta-sheet located between two globular domains consists of two beta-hairpin units and thus can be considered as a prototype of self-assembly. We constructed self-assembly mimics of different sizes and determined their atomic structures using x-ray crystallography and NMR spectroscopy. Highly regular beta-sheet geometries were maintained in these structures, and peptide units had a nearly identical conformation, supporting the concept that a peptide in the regular beta-geometry is primed for self-assembly. However, we found small but significant differences in the relative orientation between adjacent peptide units in terms of beta-sheet twist and bend, suggesting their inherent flexibility. Modeling shows how this conformational diversity, when propagated over a large number of peptide units, can lead to a substantial degree of nanoscale polymorphism of self-assemblies.
尽管富含β折叠的自组装体是多肽的一种主要结构类型，也是深入研究的焦点，但由于其不溶性和非晶态性质，人们对其原子结构和动力学知之甚少。我们开发了一种蛋白质工程策略，可在水溶性分子中捕获一个自组装片段。连接预定义数量的自组装肽单元，并封闭β折叠末端以防止聚集，从而产生单分散的可溶性蛋白质。我们通过使用伯氏疏螺旋体表面蛋白（OspA）测试了该策略，该蛋白位于两个球状结构域之间的单层β折叠由两个β发夹单元组成，因此可被视为自组装的一个原型。我们构建了不同大小的自组装模拟物，并使用X射线晶体学和核磁共振光谱确定了它们的原子结构。在这些结构中保持了高度规则的β折叠几何形状，肽单元具有几乎相同的构象，这支持了处于规则β几何形状的肽易于自组装的概念。然而，我们发现相邻肽单元之间在β折叠扭曲和弯曲方面的相对取向存在微小但显著的差异，表明它们具有内在的灵活性。建模显示，当这种构象多样性在大量肽单元中传播时，如何导致自组装体在纳米尺度上具有相当程度的多态性。
