蛇毒中毒被WHO列为“被忽视的疾病”之首，每年导致较高的伤残率，开展广谱蛇伤药物研究十分必要。我们前期在华游蛇血清中发现了广谱抗蛇毒蛋白PLIγ。最近我们发现，将超致死量的五步蛇毒或眼镜蛇毒注入华游蛇体内，蛇不死亡且中毒症状不明显，动物实验显示华游蛇肝脏提取物有较强的抗出血和肌肉毒活性。我们用自制的五步蛇毒-琼脂糖亲和层析柱对华游蛇血清进行了初步实验，发现有多个蛋白与蛇毒亲和柱结合。以上研究表明华游蛇体内存在多种蛇毒抑制剂，但其性质及作用的毒素靶点尚不清楚。本项目我们将采用组学策略，系统筛选华游蛇肝脏抗蛇毒蛋白质组，并以抗蛇毒组-亲和柱进行反向实验，利用shotgun-LC/MS对华游蛇抗毒素组及其毒素靶标进行鉴定。对新抑制剂进行抗出血毒、肌肉毒、神经毒、系统毒检测与基因克隆，并分析抑制剂基因表达与注毒量关系。本项目将建立基于毒素组的药物筛选方法，为新型抗蛇毒药物研究奠定基础。

Snake envenomation was considered as the top of neglected disease by WHO, which often leads to a high incidence of death or physical disability. The drug discovery of novel broad-spectrum antivenom is thereby urgent and meaningful. We had discovered a wide spectrum antivenom protein, namely PLIγ from the snake Sinonatrix annularis serum. We recently performed injection of supralethal dose snake venom of D.acutus or N. atra on the snake S. annularis, and found the snakes kept alive with slight envenomed symptoms. The liver lysate of S. annularis also showed strong anti-hemorrhage and anti-myonecrosis effects in vivo. We exerted binding experiment of S. annularis serum through self-prepared venom-agarose affinity column, and found a dozen of proteins were fished. These results suggested that S. annularis contains a cocktail of snake venom inhibitors with unclear properties and toxin targets. In this project, we are going to screen the antivenom proteome against D. acutus and N. atra venom from S. annularis liver lysate using “-omics” strategy, as well the reverse binding of hepatic antivenome to their toxin targets. The resulted novel inhibitors and their target toxins will be identified by shotgun LC/MS. We are going to determine the antivenom activities, including the inhibition to hemorrhage, myonecrosis, neurotoxicity and systemic effects, and clone the genes of these novel inhibitors. Furthermore, these inhibitors’ gene expression in liver of envenomed snakes will be assayed by qPCR, in aims to verify the neutrilization ability against evenoming amouts. The project will establish a venome-targeted technique for the biopanning of antidotes, which is beneficial for novel antivenom drug discovery.