Serumpeptidome解析.ppt

Identification and Separation of Proteomics 分离与鉴定蛋白质比核酸困难得 Much more difficult to separate and identify proteins than nucleic acid 对蛋白质组进行表征, 即全部蛋白质的分离、鉴定及图谱制作。 Protocol: First step: separate proteins from samples Second step: identify those proteins Identification and Separation of Proteomics 双向凝胶电泳 two dimentional electrophoresis, 2- DE 蛋白质组多维色谱分离技术 chromatographic fractionation 质谱相关的蛋白质鉴定技术 mass-spectrum 同位素亲和标签技术 isotope- coded affinity tag, ICAT 蛋白质与抗体阵列芯片的检测 protein array Identification and Separation of Proteomics 质谱相关的蛋白质鉴定技术 mass-spectrum 基本原理: 样品分子离子化后, 根据不同离子间质荷比 m/ z 的差异来分离并确定分子质量. Principle: after the sample molecules are ionized, according to the mass-to-charge ratio m / z of the differences between the different ions to separate and determine the molecular mass . Identification and Separation of Proteomics 质谱相关的蛋白质鉴定技术 mass-spectrum 分子质量 molecular mass 是一个蛋白质、多肽或氨基酸最基本的特征, 且这种特征是专一的。 Fundamental feature of a protein, peptide or AA., and it is unique. 不同的氨基酸组成、氨基酸序列、修饰方式、蛋白质间的结合方式、位点差异都可以在蛋白质的分子质量上得以体现 diversity of different amino acid composition, sequence, modification, conjugation ways, et al. leads to distinct molecular mass 质谱 mass spectrum 相关的蛋白质鉴定技术 因此, 对蛋白质以及组成蛋白质的多肽、氨基酸的分子质量测定实际上就是对蛋白质种类、结构和性质的鉴定 In fact, molecular mass determination of proteins , or peptides and amino acids composing of proteins identification of category, structure and quality of proteins Identification and Separation of Proteomics 质谱鉴定蛋白质或多肽的方式主要有两种: 肽质量指纹图分析 peptide mass fingerprinting, PMF 氨基酸序列分析 串联质谱 tandem mass spectrometry, MS/MS Identification and Separation of Proteomics 基质辅助激光解吸离子化飞行时间质谱 matrix- associated laser desorption / ionization time of flight mass spectrometry, MALDI- TOF-MS MALDI- TOF-MS 的应用, 使蛋白质鉴定的速度和灵敏度得以大大提高, 取得了革命性的突破。 The low molecular weight region of the serum peptidome contains protein fragments derived from 2 sources: a high-abundance endogenous circulating proteins and b cell and tissue proteins. While s