蛇毒神经毒素类似物的免疫学分析

The Immunological Analyses of Ten Neurotoxin-like Proteins from Snakes*

选取眼镜蛇蛇毒和银环蛇蛇毒中 1 0个差异较大的神经毒素类似物 ,通过麦芽糖融合表达系统使它们在大肠杆菌中得到高效可溶性表达 .通过 Amylose- Sepharose6B亲和树脂纯化这些融合蛋白 .分别制备这 1 0种融合蛋白多克隆抗体 .同时将这 1 0种神经毒素类似物进行硫氧还蛋白融合表达 ,表达产物基本上都是包涵体 .以硫氧还蛋白融合表达产物为抗原和上述 1 0种神经毒素类似物抗血清进行 Western blotting.结果显示 ,大多数神经毒素类似物之间没有免疫交叉反应 ,表明这些神经毒素类似物的抗原性差别较大 。

Neurotoxins from snake venom are low molecular weight polypeptides,but they are rich in disulfide bonds and possesses strong and specific physiological activities.Snake neurotoxins have been good materials to study for relationships of protein structure and function.With the development of modern molecular biology,some cDNAs for neurotoxin like proteins were cloned from the venom glands of Naja naja atra and Bugrarus multicinctus multicinctus,but biological functions of their coding proteins were not known.In order to shed light on their classification,evolution and characterization of structure and function,the immunological analyses were made to study these neurotoxin like proteins.Ten kinds of neurotoxin like proteins were efficiently expressed in E.coli as soluble MBP fusion proteins.They were purified by Amylose Sepharose 6B affinity chromatography.Ten kinds of multiclonal antibodies were prepared by using the purified fusion proteins.The ten neurotoxin like proteins were also expressed in the form of inclusion bodies as TrxA fusion proteins.The ten inclusion bodies were taken as antigens in Western blotting.The results showed that most of the neurotoxin like proteins could not cross react between themselves,indicating that they might be different in their classification and evolution.