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

先前我们报道了缺口β -糖蛋白I （b2GPI）通过其结构域V与血管抑制素结合并发挥促血管生成作用（Nakagawa等）。血2009)。本研究旨在探讨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生成。采用反flag柱或镍柱对产物进行纯化。利用表面等离子体共振评估完整/缺口b2GPI结构域V与血管抑制素4.5 （AS4.5）之间的结合。缺失的b2GPI结构域V在KD为~ 5 × 10-8M时特异性结合到AS4.5上，而完整的b2GPI结构域V没有特异性结合。使用人脐带内皮细胞（HUVECs）进行细胞增殖实验，在存在或不存在完整的/缺口的b2GPI结构域V的情况下，完整的和缺口的b2GPI结构域V在0.4uM时均能轻微抑制细胞增殖。综上所述，b2GPI的切口结构域V与AS4.5结合，在整个b2GPI中观察到切口形式。

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.