蛋白質體在生物醫學上之應用

Genomics study of molecular organization of genomes, their information content, and gene products they encode divided into three areas –structural genomics physical nature of genomes –functional genomics how genome functions –comparative genomics compares genomes of different organisms

Functional Genomics determination of how genome works three common approaches –genome annotation –study of RNA-level gene expression –study of protein-level gene expression

Genome Annotation

Nature Genetics 21:10-14, Supplement (1999)

Evaluation of Protein-Level Gene Expression proteome –entire collection of proteins that an organism produces proteomics –study of the proteome

Types of proteomic analysis functional proteomics –study of function of cellular proteins, how they interact, and ways they are regulated structural proteomics –using known protein structures to predict three-dimensional structures of other proteins and protein complexes

Evaluation of Protein-Level Gene Expression… Two-dimensional gel electrophoresis coupled with mass spectrometry (Gel-based proteomics) Two-dimensional liquid chromatography (2-D LC) coupled with mass spectrometry (Chromatography-based proteomics) SELDI  Protein chip coupled with mass spectrometry Antibody array

Two-dimensional gel electrophoresis when coupled with mass spectrometry, used to: –determine mass of each protein –determine amino acid composition or sequence of each protein –identify protein

Pandey et.al., Nature 405 (2000)

Silver stain

Sensitivity: 1-2 ng Use formaldehyde or glutaraldehyde as reductant and fixative Aldehydes form Shiff’s bases with 1  and 2  amines Not suitable for Edman degradation and mass I.D. Omit aldehydes: lost sensitivity Narrow linear range  over saturation

SYPRO Ruby Sensitivity: 1-2 ng Large linear range Speckle: mistaken as spots Metal contamination

LavaPurple Epicocconone Isolated from Epicoccum nigrum Enamine Primary amine, Lysine, Arginine,

Two-dimensional difference in gel electrophoresis