Role of recombinant nicked beta2-glycoprotein I domain V in angiogenesis

先前我们报道了NICKβ2-糖蛋白I(B2GPI)通过其结构域V与血管抑素结合并发挥促血管生成作用(Nakagawa等人)。血液2009)。本研究的目的是探讨b2GPI结构域V在血管生成中的作用。重组b2GPI结构域V在哺乳动物细胞表达系统和细菌中得到表达。用抗FLAG柱或镍柱对产物进行纯化。用表面等离子共振技术检测完整/缺失的b2GPI结构域V与血管抑素4.5(AS4.5)的结合。缺失的b2GPI结构域V在Kd为5×10-8M时与AS4.5特异性结合，而完整的b2GPI结构域V无特异性结合。人脐静脉内皮细胞(HUVECs)在存在或不存在完整/有缺口b2GPI结构域V的情况下进行细胞增殖实验，完整和有缺口的b2GPI结构域V在0.4uM时对细胞的增殖有轻微的抑制作用。

Previously we reported nicked beta2-glycoprotein I (b2GPI) binds to angiostatin via its domain V and exerts pro-angiogenic effect (Nakagawa et al. Blood 2009). The aim of this study was to investigate the role of b2GPI domain V in angiogenesis. Recombinant b2GPI domain V was generated in mammalian cell expression system and bacteria. Purification of the product was done using anti-FLAG column or nickel column. Binding between intact/nicked b2GPI domain V and angiostatin 4.5 (AS4.5) was evaluated using surface plasmon resonance. Nicked b2GPI domain V specifically bound onto AS4.5 at KD of ~ 5 x 10-8M, whereas intact b2GPI domain V showed no specific binding. Cell proliferation assay using human umbilical endothelial cells (HUVECs) were performed in the presence or absence of intact/nicked b2GPI domain V. Both intact and nicked b2GPI domain V slightly inhibited cell proliferation at 0.4uM.In conclusion, nicked b2GPI domain V bound to AS4.5, as observed in nicked form of whole b2GPI.