第三組 : 李翩翩 許睿琪 林宣辰 李宜潔 林家弘

1. 第三組: 李翩翩 許睿琪 林宣辰李宜潔 林家弘

2. Background • 1953 Simpson • Protein degradation needs ATP • 1960 discovery of lysosome • 1975 Goldstein • discovery of ubiquitin • 1977 Goldberg • Reticulocyte: protein degradation needs ATP • Ecoli : Lon enzyme

3. 2004 Nobel Prize in chemistry Aaron Ciechanover Israel Technion – Israel Institute of Technology • Avram Hershko • Israel • Technion – Israel Institute of Technology • Irwin Rose • USA • University of California

4. Process • 1971 Hershko開始專門研究能量對蛋白質分子降解的作用 • 1977 Hershko與Ciechanover著手研究 • 不含溶酶體的未成熟網織紅細胞 • 細胞萃取液中含兩個部分 • 以放射性同位素標定蛋白質

5. 1978至Fox chase癌症研究中心與Rose一同研究 • 證實其一為APF-1 • 熱穩定特性 • 分子量9000 • ATP-dependent proteolysis factor 1 • 1980 APF-1 以共價鍵與protein結合 • 多次泛素化(polyubiquitination) • 多個APF-1鍵結在同一個protein →proteasome 分解機制啟動

6. 共價結合酵素 共價結合酵素 蛋白受質 蛋白受質 proteasome proteasome • 1981~1983三人發展出一套 “多重步驟泛素標籤化假說” • E1 E2 E3三種酵素與泛素標籤化有關 E1 活化酵素 E2 攜帶酵素 E3 泛素連接酶

7. General introduction of protein degradation

8. Two major systems for intracellular protein degradation Lysosome pathway Proteasome pathway

10. Ubiquitin Tiny (76 aa) cytosolic protein 1 Carboxylterminal Gly on C-terminal Lys on protein substrate Gly on C-terminal Cys on enzyme (E1/E2) Peptide bond Thiolester bond

11. ubquitination

12. Ubiquitin-proteasome pathway’s ubiquitin enzymes E1 Ubiquitin-activating enzyme E2 Ubiquitin-carrier enzyme E3 Ubiquitin-protein ligase 活化酵素 攜帶酵素 泛素／類泛素連接酶

14. Proteasome

15. 26S proteasome泛素 目標蛋白分解複合體 ，總共分為兩部分： 19S：調節部分 （regulatory particle, RP） 1.辨認目標蛋白 2.去除泛素 3.與目標蛋白之共價結合 4.將目標蛋白解旋，送入20S 20S：蛋白分解酶 （core protease, CP） 負責目標蛋白的分解。 Proteasome

16. 19S regulator 19S：調節部分 （regulatory particle, RP） 1.辨認目標蛋白 2.去除泛素 3.與目標蛋白之共價結合 4.將目標蛋白解旋，送入20S 由一個Lid和一個Base組成，Base中含有ATPases，可以分解ATP產生能量。 19S regulator中的isopeptidase可以將ubiquitin從substrate protein上移除。

17. 20S core 20S：蛋白分解酶 （core protease, CP） 負責目標蛋白的分解。 四個環，αββα，各由7個次單元組成。 三個室（chamber）：Antechamber（前室） Catalytic chamber（催化反應室） Antechamber。 三個catalytic sites，位於某些β次單元上。

20. Relatedfunction NF-kB(nuclear factor-kappa B)&IKB

21. Cell cycle • Mechanism of E3 in cell cycle • E3 enzyme in cell cycle=APC (anaphase-promoting complex) • Importance to the separation of chromosomes

23. Cell cycle • Mechanism of E3 and p53 in apoptosis • p53:continual production and breakdown (through ubiquitination) • DNA injury  p53 is phosphorylated by ATM kinase and no longer bind E3(mdm2) • quantity of p53 risesstop the cell cycle to repair DNA damage; apotosis • Cancer :change in E3breakdown of p53  false in DNA repair & apoptosis  cancer

24. Cystic Fibrosis • common lethal autosomal recessive disease in white populations • disease of exocrine glands. • A hereditary disease that affects mainly the aspiratory and digestive system, leading to the accretion of thick phlegm in the organs which impairs their function • Excessive mucus block the tracts

25. non-functioning CFTR, "cystic fibrosis transmembrane conductance regulator". • CFTR can be found in tissues that produce mucus, sweat, saliva, tears and digestive enzymes. • genetic damage, loss of the amino acid phenylalanine in the CFTR protein • The mutation causes faulty folding of the CFTR and leads to the protein being retained in the cell's control system.

26. The incorrectly folded protein is destroyed through ubiquitin-mediated protein breakdown. • A cell with no functioning chloride channel can no longer transport chloride ions through its wall.

27. Parkinson's disease • a degenerative disorder of the central nervous system that often impairs the sufferer's motor skills and speech • Caused by the degradation of nerve cells in basal ganglia substantial nigra. • Lewy body

28. Two systems Chaperone Ubiquitin-proteasome

29. Gene mutation • α-synuclein • parkin protein

30. Thank you very much!