海藻凝集素的筛选、纯化及其性质-植物病理学专业论文.docxVIP

福建农林大学硕士学位论文摘 福建农林大学硕士学位论文 摘 要 (以抗烟草花叶病毒(Tbacco mosaic virus，TMV)、抗肿瘤 (MGc 80—31及具凝集血红细胞活性为标准，对福建沿海的10种 大型海藻(红藻6种、褐藻2种、绿藻2种)进行活性筛选。 1 0种海藻中7种具抗TMV活性，其中6种的抑制效果在60％ 以上，最高可达84．5％。9种对兔血红细胞具凝集活性，4种对 鸡血红细胞具凝集活性，且对兔血红细胞的活性要比对鸡血红 细胞高，说明海藻凝集素对兔血红细胞更敏感。l 0种海藻中多 数对胃癌MGc80-3具不同程度的抗性，其中3种抗性在50％以 上，最高的达83．2％。4 本实验以绿藻孔石莼(Ulva pertusa Kiellm)为试验材料， 经硫酸铵分级沉淀(30-80％)、牛甲状腺球蛋白．Sepharose 4B 亲和层析分离纯化得到一种凝集素(UPLl)。在PAGE及 SDS—PAGE上均显示单一蛋白染色带，测定的UPLl相对分子 量约23kD。UPLl抗TMV活性为56％；抗胃癌MGc803(1 0×) 活性为51．5％；对兔血红细胞有凝集作用。其凝血活性可被牛 甲状腺球蛋白和N一乙酰．D一氨基葡萄糖抑制，最小抑制浓度分 别为59／L，2．7659／L。UPLl 的N一端氨基酸序列为 AGITNTDNwETFAGLPLTGA，Genbank登录号为P83209。以 纯化的UPLl为抗原，制备了抗血清，效价为1：214。 (部分理化特性的研究结果表明，UPLl的分子中含有约 1．2％的中性糖；其在pH6～9范围内相对稳定；具有较高的热稳 定性，在70℃以下对兔血红细胞的凝血活性稳定，80℃时丧失 一半活性；此外，UPLl对Mnn、zn2+、M92+和ca2+等二价离 子有不同程度的依赖性。心 关键词 海藻?TMV；肿瘤；凝集素；生物活性 福建农林大学硕士学位论文Abstract 福建农林大学硕士学位论文 Abstract Ten Fujian marine algaes(six green，two red and two brown)were detected for hemagglutinins using erythrocytes of rabbit and chicken，inhibitory effect of PBS extracts on the infection of tobacco mosaic virus(TMV)on Nicotiana glutinosa and on the human stomach cancer MGc80-3．It showed that seven species inhibited the infection of TM=v(the max effect up to 84．5％、．nine species were able to agglutinate native erythrocyte of rabbit and four species were able to agglutinate native erythrocyte of chicken among ten marine algaes．The marine algae lectins were more sensitive to erythrocyte ofrabbit than to erythrocyte of chicken．Numerous algaes showed different fastness on the human stomach cancer MGc80—3．The inhibition rates of three species were higher than 50％． A new lectin，named UPLl，was isolated and purified from the green marine algae Ulva Pertusa by extraction with PBS，followed by precipitation of 30％-～80％saturation of(NH4)2S04 and an affinity chromatography on the Bovine—thyroglobulin-Sepharose 4B column．UPLl was proved to be homogeneous by PAGE．By SDS—PAGE，authour measured subunit raletive molecular mass幻be about 23kD．Z西e拍e inhibition rate o