家蚕孤雌生殖相关蛋白PGL的鉴定与生物信息学分析  

Identification and Bioinformatic Analysis of a Parthenogenesis Associated Protein Phosphatidylinositol Glycan in Bombyx mori

作　　者：张文平[1] 吕正兵[1] 龙晓辉[1] 聂作明[1] 夏佳音[1] 张耀洲[1] 

机构地区：[1]浙江理工大学生命科学学院生物化学研究所,杭州310018

出　　处：《蚕业科学》2010年第6期889-896,共8页ACTA SERICOLOGICA SINICA

基　　金：国家自然科学基金项目(No.30277056);浙江省教育厅项目(No.Y201019098);浙江省生物医学工程重中之重学科开放基金项目(No.SWYX0915)

摘　　要：应用二维凝胶电泳(2DE)技术分离有性生殖蚕卵与孤雌生殖蚕卵的差异蛋白,通过基质辅助激光解吸电离串联飞行时间质谱(MALDI-TOF-TOFMS)分析获得大量小肽的序列特征。与已构建的家蚕cDNA文库及蛋白质数据库进行比对,确定一个在家蚕孤雌生殖过程中表达量显著上调的差异蛋白为磷脂酰肌醇蛋白聚糖(phosphatidylinositol glycan,PGL)。为了探讨PGL蛋白在家蚕孤雌生殖发生中的作用,采用cDNA末端快速扩增(RACE)技术扩增目的基因5′端片段,获得PGL蛋白基因的全长cDNA,生物信息学分析显示该基因cDNA全长1038bp,编码345个氨基酸,其编码蛋白理论分子质量为39.516kD,等电点为5.845,包含1个保守的GPI8结构域,信号肽概率为0.997,可能为分泌型蛋白,预测蛋白功能域包含酪蛋白激酶Ⅱ磷酸化、N糖基化、蛋白激酶C磷酸化等多个功能位点。依据PGL蛋白的理化性质、结构功能推测其有可能在不同程度上通过细胞信号转导途径参与了家蚕孤雌生殖发生过程并发挥重要作用。Two-dimension gel electrophoresis （2DE） technique was used to identify differential protein spots between Bombyx mori eggs from normal sexual reproduction and those from parthenogenetic reproduction.After these protein spots were analyzed by MALDI-TOF-TOF MS,a large number of short peptide sequences were obtained.Alignments with sequences in a constructed cDNA library and in predicted protein database of Bombyx mori led to the identification of a phosphatidylinositol glycan protein （PGL） of which expression was up-regulated significantly in the process of parthenogenesis.In order to elucidate the function of PGL protein in Bombyx mori parthenogenesis,the full-length PGL protein gene was obtained by means of 5＇-RACE.Bioinformatic analysis showed that this gene contains 1 038 nucleotides and encodes 345 amino acids with a predicted molecular weight of 39.516 kD and pI of 5.845.Moreover,the encoded protein harbors a conserved domain GPI8.Prediction of signal peptide showed that the probability of signal peptide was 0.997,suggesting its identity as a secreted protein.Functional domain prediction indicated that the protein has various functional sites including casein kinase Ⅱ phosphorylation,N-glycosylation,and protein kinase C phosphorylation sites.According to PGL protein＇s physical and chemical properties and its structural and functional domains,it is suggested that PGL protein participates in the process of Bombyx mori parthenogenesis and plays important roles in it through cell signaling pathway.

关 键 词：家蚕孤雌生殖 差异表达蛋白 二维凝胶电泳 基质辅助激光解吸电离串联飞行时间质谱 磷脂酰肌醇蛋白聚糖 基因克隆 生物信息学分析 

分 类 号：S881.2[农业科学—特种经济动物饲养] Q51[农业科学—畜牧兽医]