【摘要】 通过磷酸缓冲液浸提、硫酸铵沉淀、DEAE-52纤维素离子交换层析等步骤,从草菇(Volvariella volvacea)菌丝体中分离得到凝集素,进而对蛇、鸭、小鼠、鸡、驴、鸽子、仓鼠7种动物的血红细胞进行凝血实验,并分别用N-溴代丁二酰亚胺(N-bromosuccinimide, NBS)、焦炭酸二乙酯(diethyl pyrocarbonate, DEPC)、苯甲基磺酰氟(phenylmethylsulfonyl fluoride, PMSF)对草菇菌丝体凝集素中的色氨酸残基、组氨酸残基、丝氨酸残基进行化学修饰,检测其凝血活性。结果表明:草菇菌丝体凝集素对蛇、鸭、小鼠、驴、鸽子、仓鼠血红细胞的凝血效价分别为2~5、2~4、2~7、2~7、2~4、2~6,对鸡血红细胞无凝血活性。色氨酸残基被修饰后凝集素凝血活性丧失,组氨酸残基、丝氨酸残基被修饰后对凝集素凝血活性无影响,表明色氨酸残基处于草菇菌丝体凝集素凝血活性中心,是维持草菇菌丝体凝集素凝血活性的必需氨基酸,而组氨酸残基、丝氨酸残基并非维持凝集素凝血活性必需的氨基酸。更多还原

【Abstract】 Using phosphate buffer extraction, ammonium sulfate precipitation and DEAE-52 cellulose ion exchange chromatography, lectin was isolated from Volvariella volvacea mycelia. The obtained lectin was subjected to coagulation tests on red blood cells of snake, duck, mouse, chicken, donkey, pigeon and hamster, respectively. Then the tryptophan, histidine and serine residues of the lectin were chemically modified with N-bromosuccinimide(NBS), diethyl pyrocarbonate(DEPC) and phenylmethylsulfonyl fluoride(PMSF) respectively and then tested for coagulation activity. The results showed that the agglutination titers of V. volvacea mycelium lectin for snake, duck, mouse, chicken, donkey, pigeon and hamster red blood cells were 2~5, 2~4, 2~7,0, 2~7, 2~4 and 2~6, respectively. V. volvacea mycelium lectin had no coagulation activity on chicken red blood cells. NBS modification on tryptophan resulted in loss of coagulation activity, whereas modifications on histidine and serine had no effect on coagulation activity. The results suggested that tryptophan was at the active center of the lectin to provide coagulation activity, whereas histidine and serine were not essential amino acids to maintain the coagulation activity of the lectin.更多还原