桶形芋螺毒液蛋白质组分析及二硫键异构酶的鉴定  

TWO-DIMENSIONAL ELECTROPHORESIS ANALYSIS OF CONUS BETULINUS LINNAEUS VENOM PROTEOME AND IDENTIFICATION ON PROTEIN DISULFIDE ISOMERASE

作　　者：彭超[1,2,3] 张容鹄[1,2] 高炳淼[1,2] 长孙东亭[1,2] 张本[1,2] 罗素兰[1,2] 

机构地区：[1]海南大学热带生物资源教育部重点实验室,海口570228 [2]海南大学海口市海洋药物重点实验室,海口570228 [3]深圳华大基因研究院,深圳518083

出　　处：《水生生物学报》2012年第6期1063-1070,共8页Acta Hydrobiologica Sinica

基　　金：国家自然科学基金(81160503);国家国际科技合作专项(2011DFR31210);国家863计划(2012AA021706);长江学者和创新团队发展计划(IRT1123)资助

摘　　要：蛋白质二硫键异构酶(PDI)是内质网新生肽链折叠中一个重要的折叠酶。在热带药用海洋生物芋螺的毒液中,富含PDI酶,该酶对于毒液中芋螺毒素神经肽的体内氧化折叠至关重要。研究上样量、水化方式与时间、除盐步骤、聚焦电压和时间、平衡时间、凝胶电泳方法和染色方法等方面,优化并建立了较为理想的芋螺毒液蛋白样品双向电泳的实验条件与方法;通过双向电泳和MALDI-TOF-MS质谱分析技术,从海南产桶形芋螺(Conus betulinus Linnaeus)毒液蛋白中成功分离鉴定出PDI等9种蛋白;通过双向电泳和PDQuest软件分析了并比较了三种不同大小桶形芋螺毒液总蛋白的差异性,并质谱鉴定出5个明显的差异蛋白点。研究建立了桶形芋螺毒液蛋白双向电泳分析及质谱鉴定的技术方法,为后续大量分离纯化出天然的芋螺PDI酶以及利用蛋白质组学技术方法深入研究芋螺毒液特征提供了重要的基础。Conopeptides is one kind of small precious nerve toxin peptides, which is excreted from Conus snails. Be cause the post translational processing of conopeptides is complicated, it is very difficult to synthesize conopeptides peptide with biological activity by the method named chemical synthesis. Many proteins from Conus snails, such as protein disulfide isomerase （PDI）, prolylhydroxylase, proline isomerase, and so on, which promote the synthesis and activity formation of conopeptides. The protein disulfide isomerase （PDI） is an important foldase in folding of the nas- cent polypeptide chain of the endoplasmic reticulum （ER）. The venom of tropic officinal marine cone snails is abundant with PDI which is important in assisting the folding and maturation of secretory conopeptides in vivo. As an effective and direct approach of research in all proteins, proteomics represents a novel methodological way to investigate the expression of all proteins of a cell, body fluid or organism, as well as to analyze their structure, function, interaction and modification. Proteomics has been widely applied in studying regulation of metabolism, genetics and development, physiology and biochemistry, pharmacology and toxicology. And many technologies of proteomics were available to research Conus snails venom protein in integer protein level. The report was to analyze the total proteins by the widely use technologies including two-dimensional eleetrophoresis （2-DE）, mass spectrum, chromatogram analysis and biology informations, so as to lay a good foundation for further research on Proteomics of Conus venom. Previous trials on dif- ferent electrophoresis methods showed that SDS-PAGE electrophoresis is difficult to detect protein with low molecular weight, while Trieine-SDS-PAGE electrophoresis can detect low molecular weight protein clearly. But in the 2-DE ex- periment, the Tricine-SDS-PAGE electrophoresis makes protein point deform and gets a blurry 2-DE map for taking more time than SDS-PAGE electrophoresis. So in

关 键 词：蛋白质二硫键异构酶 芋螺毒素 双向电泳 毒液蛋白组学 

分 类 号：Q512[生物学—生物化学]