| ○種別 (必須): | □ | 学術論文 (審査論文)
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| ○言語 (必須): | □ | 英語
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| ○カテゴリ (推奨): |
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| ○学究種別 (推奨): |
| ○組織 (推奨): |
| ○著者 (必須): | 1. | (英) Kameshita Isamu (日) (読)
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| 2. | (英) Yamashita Sho (日) (読)
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| 3. | 片山 将一 ([徳島大学.大学院医歯薬学研究部.薬学域.先端薬学教育研究プロジェクト]/[徳島大学.大学院医歯薬学研究部.薬学域.薬科学部門.生命薬学系.医薬品病態生化学])
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| 4. | (英) Senga Yukako (日) (読)
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| 5. | (英) Sueyoshi Noriyuki (日) (読)
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| ○題名 (必須): | □ | (英) TandeMBP: generation of a unique protein substrate for protein kinase assays. (日)
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| ○副題 (任意): |
| ○要約 (任意): | □ | (英) Myelin basic protein (MBP) is one of the major components of central nervous system myelin and has multiple sites for protein phosphorylation. Therefore, it has been widely used as a substrate for in vitro assays of various protein kinases. In this study, to obtain more efficient substrates for protein kinase assays than commercially available MBP from bovine brain, we produced various recombinant MBPs using Escherichia coli expression systems. Three splice isoforms of mouse MBP were expressed in E. coli and successfully purified using a new protocol consisting of HCl extraction, urea treatment and affinity purification with HiTrap Chelating HP column. The recombinant MBP isoforms thus obtained served as more efficient substrates for protein kinases than MBP isolated from bovine brain. To generate an even better substrate for protein kinase assays, we produced a hybrid protein composed of two different MBP isoforms connected in tandem, designated TandeMBP. TandeMBP was readily expressed in E. coli and could be purified by the newly developed simple procedure. TandeMBP was phosphorylated by various Ser/Thr protein kinases more efficiently than the other MBP isoforms. Taken together, TandeMBP will become a powerful tool for in vitro assays to analyse various protein kinase activities. (日)
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| ○キーワード (推奨): | 1. | (英) Amino Acid Sequence (日) (読)
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| 2. | (英) Animals (日) (読)
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| 3. | (英) Blotting, Western (日) (読)
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| 4. | (英) Cattle (日) (読)
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| 5. | (英) Enzyme Assays (日) (読)
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| 6. | (英) Molecular Sequence Data (日) (読)
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| 7. | (英) Myelin Basic Protein (日) (読)
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| 8. | (英) Phosphorylation (日) (読)
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| 9. | (英) Protein Isoforms (日) (読)
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| 10. | (英) Protein Kinases (日) (読)
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| 11. | (英) Recombinant Fusion Proteins (日) (読)
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| 12. | (英) Reproducibility of Results (日) (読)
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| 13. | (英) Sequence Homology, Amino Acid (日) (読)
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| 14. | (英) Substrate Specificity (日) (読)
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| ○発行所 (推奨): |
| ○誌名 (必須): | □ | The Journal of Biochemistry ([日本生化学会])
(pISSN: 0021-924X, eISSN: 1756-2651)
| ○ISSN (任意): | □ | 1756-2651
ISSN: 0021-924X
(pISSN: 0021-924X, eISSN: 1756-2651) Title: Journal of biochemistryTitle(ISO): J BiochemSupplier: Oxford University PressPublisher: Oxford University Press (NLM Catalog)
(医中誌Web)
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| ○巻 (必須): | □ | 156
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| ○号 (必須): | □ | 3
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| ○頁 (必須): | □ | 147 154
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| ○都市 (任意): |
| ○年月日 (必須): | □ | 西暦 2014年 4月 8日 (平成 26年 4月 8日)
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| ○URL (任意): |
| ○DOI (任意): | □ | 10.1093/jb/mvu025 (→Scopusで検索)
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| ○PMID (任意): | □ | 24713852 (→Scopusで検索)
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| ○備考 (任意): | 1. | (英) PublicationType: Journal Article (日)
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| 2. | (英) PublicationType: Research Support, Non-U.S. Gov't (日)
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