Chakrabarty D, Bhattacharyya D, Sarkar H S, Lahiri S C
Indian Institute of Chemical Biology, Calcutta.
Toxicon. 1993 Dec;31(12):1601-14. doi: 10.1016/0041-0101(93)90344-i.
A haemorrhagic toxin (VRH-1) has been purified to homogeneity from Vipera russelli russelli venom by subjecting it to chromatography twice successively on CM-Sephadex C-50. It is a protein of mol. wt 22,000 and contains one mole of Mg2+. Intradermal administration of this haemorrhagin in mice resulted in severe lung haemorrhage but produced little haemorrhage in skin. This apparent organ preference led us to develop a new haemorrhage assay method utilizing dye diffusion from lung in vitro. Proteolytic activity of VRH-1 was demonstrated using dimethylcasein as substrate following quantitation by reaction with trinitrobenzoyl sulfonic acid. Both haemorrhagic and proteolytic activities of VRH-1 were inhibited by serine protease inhibitors like phenylmethyl sulfonyl fluoride and chymostatin, but metal chelators had no effect. Lung haemorrhage is unlikely to be a direct reflection of a high local concentration of VRH-1. The administration of supernatant generated by incubation of chopped liver from untreated mouse and VRH-1 (in subhaemorrhagic dose) results in severe lung haemorrhage. This raises the possibility that VRH-1 leads to the formation of intermediate(s) which causes the haemorrhage.
通过先后两次在CM - Sephadex C - 50上进行层析，从圆斑蝰蛇毒中纯化出一种出血毒素（VRH - 1），使其达到同质。它是一种分子量为22,000的蛋白质，含有一摩尔Mg2 +。给小鼠皮内注射这种出血毒素会导致严重的肺出血，但在皮肤上产生的出血很少。这种明显的器官偏好促使我们开发一种利用染料在体外从肺扩散的新出血检测方法。以二甲基酪蛋白为底物，通过与三硝基苯甲酰磺酸反应进行定量后，证明了VRH - 1的蛋白水解活性。VRH - 1的出血和蛋白水解活性均受到丝氨酸蛋白酶抑制剂如苯甲基磺酰氟和抑肽酶的抑制，但金属螯合剂没有作用。肺出血不太可能直接反映VRH - 1的高局部浓度。用未处理小鼠的切碎肝脏与VRH - 1（亚出血剂量）孵育产生的上清液给药会导致严重的肺出血。这增加了VRH - 1导致形成引起出血的中间产物的可能性。
