Pedersen Dennis Vestergaard, Revel Margot, Gadeberg Trine Amalie Fogh, Andersen Gregers Rom
Department of Molecular Biology and Genetics, Aarhus University, Gustav Wiedsvej 10C, DK-8000 Aarhus, Denmark.
Acta Crystallogr F Struct Biol Commun. 2019 Feb 1;75(Pt 2):0. doi: 10.1107/S2053230X18018150. Epub 2019 Jan 23.
The 54 kDa protein properdin, also known as factor P (FP), plays a major role in the complement system through the stabilization of the alternative pathway convertases. FP circulates in the blood as cyclic dimers, trimers and tetramers, and this heterogeneity challenges detailed structural insight into the mechanism of convertase stabilization by FP. Here, the generation of an intact FP monomer and a variant monomer with the third thrombospondin repeat liberated is described. Both FP monomers were excised from recombinant full-length FP containing internal cleavage sites for TEV protease. These FP monomers could be crystallized, and complete data sets extending to 2.8 Å resolution for the intact FP monomer and to 3.5 Å resolution for the truncated variant were collected. The principle of specific monomer excision and domain removal by the insertion of a protease cleavage site may be broadly applicable to structural studies of oligomeric, flexible and modular proteins.
54千道尔顿的备解素蛋白，也称为因子P（FP），通过稳定替代途径转化酶在补体系统中发挥主要作用。FP以环状二聚体、三聚体和四聚体的形式在血液中循环，这种异质性对深入了解FP稳定转化酶的机制提出了挑战。在此，描述了完整FP单体和释放出第三个血小板反应蛋白重复序列的变体单体的产生。两种FP单体均从含有TEV蛋白酶内部切割位点的重组全长FP中切除。这些FP单体可以结晶，并收集了完整FP单体分辨率高达2.8埃、截短变体分辨率高达3.5埃的完整数据集。通过插入蛋白酶切割位点进行特异性单体切除和结构域去除的原理可能广泛适用于寡聚、柔性和模块化蛋白质的结构研究。
