2006 Fiscal Year Final Research Report Summary
Production of monoclonal antibody to detect a wide variety of protein kinases and its application to proteome research
| Project/Area Number |
17550157
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Chemistry related to living body
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| Research Institution | Kagawa University |
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Principal Investigator |
KAMESHITA Isamu Kagawa University, Faculty of Agriculture, Professor, 農学部, 教授 (60127941)
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| Co-Investigator(Kenkyū-buntansha) |
SUEYOSHI Noriyuki Kagawa University, Faculty of Agriculture, Assistant Professor, 農学部, 助教授 (90346635)
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| Project Period (FY) |
2005 – 2006
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| Keywords | Protein kinase / Proteome / Monoclonal antibody / Biotechnology / Signal transduction |
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| Research Abstract |
1, roduction of a monoclonal antibody to detect a variety of tyrosine kinases:
In our previous study, we obtained monoclonal antibodies to detect various protein Ser/Thr kinases (Multi-STK antibodies). In this study, we attempted to produce a monoclonal antibody directed to highly conserved region of protein Tyr kinases (PTK), and obtained YK34 antibody that detected a wide variety of PTKs (Multi-PTK antibody). Multi-PTK antibody detected many of PTKs including Src family kinases, but it did not react with STK, suggesting that PTK-directed antibody was obtained.
2, expression profile of PKs in various cells as analyzed by Multi-PK antibodies:
HL-60 cells were treated with TPA to induce differentiation. Protein kinases expressed before and after differentiation were separated by two-dimensional electrophoresis and detected by Western blotting with Multi-PK antibodies. We found that several PTKs were significantly induced after TPA treatment, while some of PTKs were diminished after differentiation. We also found that expression of two unique protein kinases of 60 kDa and 200 kDa were significantly changed in INS-1 cells after treatment with high glucose. Expressions of these STKs were closely correlated with insulin secretion and thought to be correlated with diabetes.
3, Screening of novel STKs in zebrafish and mushroom:
When expression screening was carried out using cDNA libraries from zebrafish embryo and mycelia of mushroom, nearly ten novel STKs were isolated. These STKs were found to be highly expressed especially in the early stages of development, suggesting that these kinases may play crucial roles in their development. Some of these protein kinases were expressed in E.coli and characterized.
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