TY  - GEN
T1  - Application of chemical proteomics to enrich and characterize the serine/threonine phosphopeptides/phosphoproteins
A1  - Chao-Hsiung Lin
A2  - Chao-Hsiung Lin
LA  - en_US
YR  - 2005
UL  - http://buscaintegrada.pucsp.br/vufind/Record/NDLTD-TW-093YM005107010
AB  - 碩士 === 國立陽明大學 === 生物化學研究所 === 93 === Phosphorylation is among the most common and important protein modifications which are crucial for protein functions and regulations in cellular processes. Currently global analysis of serine/threonine phosphorylation remains largely unexplored due to lack of direct analysis tools. A number of chemical modification techniques were recently reported to selectively enrich serine/threonine phosphorylated proteins. In my work, application of on immobilized metal affinity chromatographyβ-elimination/aminoethanethiol addition to enrich phosphopeptides from peptides mixtures and determine the definite sites of phosphorylation are established. Through this approach, the phosphorylation sites of synthetic phosphopeptide and ��-casein are definitely identified.  In addition, the sites of p53 and Pum2H subjected to in-vitro phosphorylation by aurora kinase A are also identified. This approach can be a tool to enrich the phosphopeptides from peptide mixture and definitely determine the serine/threonine phosphorylation site. Alternatively,  β-elimination/biotin tagging can be combined with the two-dimensional gel electrophoresis to enrich and characterize the serine/threonine phosphoproteins. The serine/threonine phosphorylation profiles of RK3E-RCAN and v-Src transformed RK3E cells were established through this approach. Several proteins with differential phosphorylation status related to v-Src-induced transformation were further identified by mass spectrometric analysis. Totally, 15 differential expressed proteins in 2D-gel were identified and several of them have known serine/threonine phosphorylation site.   Among the identified differential expressed proteins, Annexin I is a known substrate of Src. The function of Annexin I is related to its serine, threonine, and tyrosine phosphorylation sites. Through the chemical tagging approach, the serine/threonine phosphoproteins can be enriched and characterized. This information gives us clues to that which proteins were subjected to the serine/threonine phosphorylation regulation. Further studies are needed to reveal the function of individual protein and the effects of its phosphorylation in the v-Src transformed cells.

ER  -