1. The Nobel Prize in chemistry 2004 Ubiquitin(Ub) 泛素 Class B Group 3 黃恒橋、劉國正、黃偲媁 黃竣聖、簡農軒、楊昆霖

2. Nobel Laureate Israel Aaron Ciechanover Israel Avram Hershko USA Irwin Rose http://big5.cctv.com/science/20041008/101027.shtml

3. The Discovery of The Ubiquitin  simple protein-degrading enzymes Trypsin Lysosome do not require energy Traditional concept

4. Paradox  1950s － the breakdown of the cell's proteins require energy  The inhibitor of lysosome has no effect on some protein degradation  Rabbit reticulocytes ( 網織紅血球 ) degrade abnormal hemoglobin

5. The Discovery of The Ubiquitin  1975 － isolated Ub from sweetbread ( 小牛胸腺 )  1977 － an extract from reticulocytes  1970s ～ 1980s － the extract could be divided into two fractions Fxn1 Fxn2

6. The Discovery of The Ubiquitin  ATP-dependent proteolysis factor 1 (APF-1)  1981 ～ 1983 － enzyme systems that binds ubiquitin to target proteins  multistep ubiquitin-tagging hypothesis

7.  Poly Ub : Ub-proteasome pathway For protein (peptide) recycling Decomposition( proteasome ) The kiss of death  Mono Ub: non-degradation Modification(methylation of Histone) The Function of Ubiquitin Target protein Ub Target protein Ub

8. The structure of Ubiquitin  A 76 a. a. protein  Stable: O H  Highly conserved  Exist in archeabacteria and all eukaryotes http://www.nottingham.ac.uk/biochemcourses/students/ub/ubindex.html

9. Binding  target protein Ub’s C-terminus the target protein’s Lysine Form isopeptide target protein Lysine Ub

10. Poly-ubiquitinization  Ub’s C-terminus Ub’s lysine  Purpose: recognize diversity protein Protein 1 Protein 2 Ub

11. Target protein L48 vs.L63 Ub L48 L63 Degraded by proteasome Degraded by lysosome

12.  Met1-Gln2-Ile3-Phe4-Val5-Lys6-Thr7-Leu8- Thr9-Gly10-Lys11-Thr12-Ile13-Thr14- Leu15-Glu16-Val17-Glu18-Pro19-Ser20- Asp21-Thr22-Ile23-Glu24-Asn25-Val26- Lys27-Ala28-Lys29-Ile30-Gln31-Asp32- Lys33-Glu34-Gly35-Ile36-Pro37-Pro38- Asp39-Gln40-Gln41-Arg42-Leu43-Ile44- Phe45-Ala46-Gly47-Lys48-Gln49-Leu50- Glu51-Asp52-Gly53-Arg54-Thr55-Leu56- Ser57-Asp58-Tyr59-Asn60-Ile61-Gln62- Lys63-Glu64-Ser65-Thr66-Leu67-His68- Leu69-Val70-Leu71-Arg72-Leu73-Arg74- Gly75-Gly76 Other lysine site(human’s Ub )

13. Ub-Proteasome Pathway http://www.bio-pro.de/imperia/md/images/grafiken/ubisystem_338x398.jpg

14. Ubiquitnation  Remarkable features The specificity of protein tagging is mainly determined by E2, E3  steps E1:Ub-activating enzyme E2:Ub-conjugating enzyme E3:Ub-ligase E2

15. E1:Ub-activating enzyme E1 AMP Ub S-H ATP Ub adenylate Cysteine Adenylation C-terminus Cysteine Activation

16. E2:Ub-conjugating enzyme E2 AMP Ub adenylate Ub E1 S-HNew E3 1.Connection of polyUb 2.Elongation of Ub with E3

17. E3:Ub-ligase  Two types E3s which do form thiol esters with ubiquitin E3s which do not form thiol esters with ubiquitin. E3 From:www.wormbook.org/.../ubiquitinpathways.htmlwww.wormbook.org/.../ubiquitinpathways.html Recognition of the substrate

18. Proteasome  Barrel-like structure  Degrade ubiquinated protein  Use the energy of ATP

19. Structure  Core protease particle (CP) Catalytic area  Regulatory particle (RP) Ubquitin recognition Deubiquitin ATPase http://www.bioscience.utah.edu/mb/mbFaculty/hill/hill.html α β β α

20. http://www.bio-pro.de/imperia/md/images/grafiken/ubisystem_338x398.jpg

21. Ubiquitin and p53  Tumor suppressor gene  DNA repair  Apoptosis

23. Cancer reason ： HPV  Human Papillomavirus  Activate E6-AP(E3) to ubiquitinate p53  Few P53  Dysplasia

24. Medicine  LDP-341 ( PS-341)  Proscript invented in 1995  Inhibit ubiquitin  function Retain cyclin Inhibit NF-kappa B Protein pressure

25. Retain cyclin S G1 G2 cdk cyclin M BACK

26. Inhibit NF-kappa B  Promote cancer cells and vessels  Restrain apoptosis  inhibitor I kappa-B DNA I kappa B NF-kappa B BACK

27. Protein pressure  Too much protein in cell  Short time pressure  Take in interval  Disadvantage and improvement Non-selective E3 influence BACK

28. Spinocerebellar Atrophy ( 小腦萎縮症 )  Background detail Cause of disease- gene Symptom  nervous system: cell death  Behavior: paralysis  One of causes- ” inclusion bodies “( 包涵體 )  Proteins are combined by SUMO-1  SUMO(Small ubiquitin-like modifier)

29. SUMO( 類泛素 )  Small ubiquitin-like modifier  Function: Modification Proteins linked by Ub or SUMO could be recognized by cell and be sent to different organelles.  Threat

30. Abnormal SUMO In Spinocerebellar Atrophy Abnormal protein SUMO Ub Abnormal protein SUMO Abnormal protein SUMO Abnormal protein SUMO Abnormal protein SUMO Finally cell death

31. Parkinson’s Disease ( 帕金森氏症 ) Background detail Cause of disease- gene Symptom  Nervous system: death of basal ganglia ( 基底核 )and substantia nigra ( 黑質 ) cell  Behavior: tremor ； rigidity Dopamine( 多巴胺 )  Reason (related to ubiquitin) Abnormal α-synuclein gene Abnormal parkin(E3) gene

32. Abnormal α-synuclein protein Ub Abnormal α-synuclein gene Abnormal parkin(E3) gene Finally cell death Abnormal parkin(E3) Abnormal protein Abnormal α-synuclein protein Ub Abnormal α-synuclein protein Ub Abnormal α-synuclein protein Ub E2 Ub

33. Take Home Messages  Ub target substrates for degradation, traffic, or modification. The well-known function is involving ubiquitin proteasome system(UPS).  Ubiquitination has to be aided by E1:activating Ub E2:regulating poly-ubiquitinization E3:recognition of substrates

34. Take Home Messages  Mechanism of P53,HPV, Spinocerebellar Atrophy and Parkinson’s Disease are all related to the ubquitination, especially the function of E3.