A POTENT ANTIHEMORRHAGIN IN THE SERUM OF THE NONPOISONOUS WATER SNAKE NATRIX TESSELLATA - ISOLATION, CHARACTERIZATION AND MECHANISM OF NEUTRALIZATION
A POTENT ANTIHEMORRHAGIN IN THE SERUM OF THE NONPOISONOUS WATER SNAKE NATRIX TESSELLATA - ISOLATION, CHARACTERIZATION AND MECHANISM OF NEUTRALIZATION
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DOI：
10.1016/0304-4165(94)90080-9
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发表时间：
1994-12-15
影响因子：
3
通讯作者：
OVADIA, M
中科院分区：
文献类型：
--
作者：
BORKOW, G;GUTIERREZ, JM;OVADIA, M
本文报道了用35- 55%硫酸铵沉淀、SephadexG-75凝胶过滤、DEAE-Sepharose和CM-Sepharose离子交换层析及Phenyl-Sepharose疏水层析等方法，从无毒的斑腹蛇（Bothrops asper）蛇毒血清中分离得到的主要天然抗出血因子（NtAH）。纯化的蛋白质显示一条带，等电点为4.5，分子量约为880 kDa。抗出血活性在pH 5.5-11.7和高达50 ℃之间稳定，但在60 ℃下20分钟后失去活性。它不与Bothrops asper蛇毒（BaH 1）的主要沉淀素形成沉淀线，也不与整个蛇毒形成沉淀线，这表明抗出血因子不是免疫球蛋白。分离的抗出血因子NtAH的中和机制不包括出血毒素BaH 1的消化。用~（125）I标记的BaH_1毒素对NtAH进行层析和ELISA实验，证明无毒水蛇天然NtAH与棘蛇蛇毒主要毒素形成一种无活性的可溶性复合物。
The main natural antihemorrhagic factor (NtAH), which inhibits the hemorrhagic activity of Bothrops asper snake venom, was isolated from the serum of the non-poisonous water snake Natrix tessellata by ammonium sulfate precipitation at 35-55%, Sephadex G-75 gel filtration, ion exchange chromatography on DEAE-Sepharose and CM-Sepharose and hydrophobic Phenyl-Sepharose chromatography. The purified protein showed one band with an isoelectric point of 4.5 and a molecular mass of about 880 kDa. The antihemorrhagic activity was stable between pH 5.5-11.7 and up to 50 degrees C, but lost activity after 20 min at 60 degrees C. It did not form a precipitin line with the main hemorrhagin of Bothrops asper snake venom (BaH1), nor with the whole venom, which suggests that the antihemorrhagic factor is not an immunoglobulin. The mechanism of neutralization by the isolated antihemorrhagic factor NtAH did not include digestion of the hemorrhagic toxin BaH1. Chromatography of NtAH with active I-125-Iabeled BaH1 toxin as well as ELISA experiments demonstrated that the mechanism of neutralization involves formation of an inactive soluble complex between the natural NtAH of the non-poisonous water snake and the main hemorrhagin of Bothrops asper venom.