A Tale of Two Giant Proteases

  • B. Rockel
  • W. Baumeister
Conference paper
Part of the Ernst Schering Foundation Symposium Proceedings book series (SCHERING FOUND, volume 2008/1)


The 26S proteasome and tripeptidyl peptidase II (TPPII) are two exceptionally large eukaryotic protein complexes involved in intracellular proteolysis, where they exert their function sequentially: the proteasome, a multisubunit complex of 2.5 MDa, acts at the downstream end of the ubiquitin pathway and degrades ubiquitinylated proteins into small oligopeptides. Such oligopeptides are substrates for TPPII, a 6-MDa homooligomer, which releases tripeptides from their free N-terminus. Both 26S and TPPII are very fragile complexes refractory to crystallization and in their fully assembled native form have been visualized only by electron microscopy. Here, we will discuss the structural features of the two complexes and their functional implications.


Intracellular Proteolysis Tripeptidyl Peptidase Ubiquitinylated Protein Proteolytic Core Proteolytic Chamber 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



We thank J. Peters for critically reading the manuscript.


  1. Babbitt SE, Kiss A, Deffenbaugh AE, Chang YH, Bailly E, Erdjument-Bromage H, Tempst P, Buranda T, Sklar LA, Baumler J, Gogol E, Skowyra D (2005) ATP hydrolysis-dependent disassembly of the 26S proteasome is part of the catalytic cycle. Cell 121:553–565CrossRefPubMedGoogle Scholar
  2. Balow RM, Ragnarsson U, Zetterqvist O (1983) Tripeptidyl aminopeptidase in the extralysosomal fraction of rat liver. J Biol Chem 258:11622–11628PubMedGoogle Scholar
  3. Baumeister W, Walz J, Zühl F, Seemüller E (1998) The proteasome: paradigm of a self-compartmentalizing protease. Cell 92:367–380CrossRefPubMedGoogle Scholar
  4. Bosch J, Tamura T, Bourenkov G, Baumeister W, Essen LO (2001) Purification, crystallization, and preliminary X-ray diffraction analysis of the tricorn protease hexamer from Thermoplasma acidophilum. J Struct Biol 134:83–87CrossRefPubMedGoogle Scholar
  5. Brandstetter H, Kim JS, Groll M, Huber R (2001) Crystal structure of the tricorn protease reveals a protein disassembly line. Nature 414:466–470CrossRefPubMedGoogle Scholar
  6. Braun BC, Glickman M, Kraft R, Dahlmann B, Kloetzel PM, Finley D, Schmidt M (1999) The base of the proteasome regulatory particle exhibits chaperone-like activity. Nat Cell Biol 1:221–226CrossRefPubMedGoogle Scholar
  7. Burri L, Servis C, Chapatte L, Levy F (2002) A recyclable assay to analyze the NH2-terminal trimming of antigenic peptide precursors. Protein Expr Purif 26:19–27CrossRefPubMedGoogle Scholar
  8. Cascio P, Call M, Petre BM, Walz T, Goldberg AL (2002) Properties of the hybrid form of the 26S proteasome containing both 19S and PA28 complexes. EMBO J 21:2636–2645CrossRefPubMedGoogle Scholar
  9. Chand A, Wyke SM, Tisdale MJ (2005) Effect of cancer cachexia on the activity of tripeptidyl-peptidase II in skeletal muscle. Cancer Lett 218:215–222CrossRefPubMedGoogle Scholar
  10. Chen CA, Huang CX, Chen CH, Liang J, Lin WB, Ke GF, Zhang HX, Wang B, Huang JA, Han ZG, Ma LX, Huo KK, Yang XM, Yang PY, He FC, Tao T (2008) Subunit–subunit interactions in the human 26S proteasome. Proteomics 8:508–520CrossRefPubMedGoogle Scholar
  11. Ciechanover A (2005) Proteolysis: from the lysosome to ubiquitin and the proteasome. Nat Rev Mol Cell Biol 6:79–86CrossRefPubMedGoogle Scholar
  12. Davy A, Bello P, Thierry-Mieg N, Vaglio P, Hitti J, Doucette-Stamm L, Thierry-Mieg D, Reboul J, Boulton S, Walhout AJM, Coux O, Vidal M (2001) A protein–protein interaction map of the Caenorhabditis elegans 26S proteasome. EMBO Rep 2:821–828CrossRefPubMedGoogle Scholar
  13. De Winter H, Breslin H, Miskowski T, Kavash R, Somers M (2005) Inhibitor-based validation of a homology model of the active-site of tripeptidyl peptidase II. J Mol Graph Model 23:409–418CrossRefPubMedGoogle Scholar
  14. Ferrell K, Wilkinson CRM, Dubiel W, Gordon C (2000) Regulatory subunit interactions of the 26S proteasome, a complex problem. Trends Biochem Sci 25:83–88CrossRefPubMedGoogle Scholar
  15. Firat E, Huai J, Saveanu L, Gaedicke S, Aichele P, Eichmann K, van Endert P, Niedermann G (2007) Analysis of direct and cross-presentation of antigens in TPPII knockout mice. J Immunol 179:8137–8145PubMedGoogle Scholar
  16. Geier E, Pfeifer G, Wilm M, Lucchiari-Hartz M, Baumeister W, Eichmann K, Niedermann G (1999) A giant protease with potential to substitute for some functions of the proteasome. Science 283:978–981CrossRefPubMedGoogle Scholar
  17. Glas R, Bogyo M, McMaster JS, Gaczynska M, Ploegh HL (1998) A proteolytic system that compensates for loss of proteasome function. Nature 392:618–622CrossRefPubMedGoogle Scholar
  18. Glickman MH, Raveh D (2005) Proteasome plasticity. FEBS Lett 579:3214–3223CrossRefPubMedGoogle Scholar
  19. Glickman MH, Rubin DM, Coux O, Wefes I, Pfeifer G, Cjeka Z, Baumeister W, Fried VA, Finley D (1998) A subcomplex of the proteasome regulatory particle required for ubiquitin-conjugate degradation and related to the COP9-signalosome and eIF3. Cell 94:615–623CrossRefPubMedGoogle Scholar
  20. Groll M, Ditzel L, Löwe J, Stock D, Bochtler M, Bartunik HD, Huber R (1997) Structure of 20S proteasome from yeast at 2.4 angstrom resolution. Nature 386:463–471CrossRefPubMedGoogle Scholar
  21. Groll M, Brandstetter H, Bartunik HD, Bourenkov G, Huber R (2003) Investigations on the maturation and regulation of archaebacterial proteasomes. J Mol Biol 327:75–83CrossRefPubMedGoogle Scholar
  22. Hanna J, Finley D (2007) A proteasome for all occasions. FEBS Lett 581:2854–2861CrossRefPubMedGoogle Scholar
  23. Hasselgren P-O, Wray C, Mammen J (2002) Molecular regulation of muscle cachexia: it may be more than the proteasome. Biochem Biophys Res Commun 290:1–10CrossRefPubMedGoogle Scholar
  24. Hilbi H, Jozsa E, Tomkinson B (2002) Identification of the catalytic triad in tripeptidyl-peptidase II through site-directed mutagenesis. Biochim Biophys Acta 1601:149–154PubMedGoogle Scholar
  25. Hölzl H, Kapelari B, Kellermann J, Seemüller E, Sumegi M, Udvardy A, Medalia O, Sperling J, Müller SA, Engel A, Baumeister W (2000) The regulatory complex of Drosophila melanogaster 26S proteasomes: subunit composition and localization of a deubiquitylating enzyme. J Cell Biol 150:119–129CrossRefPubMedGoogle Scholar
  26. Hong X, Lei L, Kunert B, Naredla R, Applequist SE, Grandien A, Glas R (2007) Tripeptidyl-peptidase II controls DNA damage responses and in vivo gamma-irradiation resistance of tumors. Cancer Res 67:7165–7174CrossRefPubMedGoogle Scholar
  27. Huai J, Firat E, Nil A, Million D, Gaedicke S, Kanzler B, Freudenberg M, van Endert P, Kohler G, Pahl HL, Aichele P, Eichmann K, Niedermann G (2008) Activation of cellular death programs associated with immunosenescence-like phenotype in TPPII knockout mice. Proc Natl Acad Sci U S A 105:5177–5182CrossRefPubMedGoogle Scholar
  28. Ishikawa T, Maurizi MR, Steven AC (2004) The N-terminal substrate-binding domain of ClpA unfoldase is highly mobile and extends axially from the distal surface of ClpAP protease. J Struct Biol 146:180–188CrossRefPubMedGoogle Scholar
  29. Iwanczyk J, Sadre-Bazzaz K, Ferrell K, Kondrashkina E, Formosa T, Hill CP, Ortega J (2006) Structure of the Blm10–20S proteasome complex by cryo-electron microscopy. Insights into the mechanism of activation of mature yeast proteasomes. J Mol Biol 363:648–659CrossRefPubMedGoogle Scholar
  30. Kisselev AF, Akopian TN, Woo KM, Goldberg AL (1999) The sizes of peptides generated from protein by mammalian 26 and 20 S proteasomes—implications for understanding the degradative mechanism and antigen presentation. J Biol Chem 274:3363–3371CrossRefPubMedGoogle Scholar
  31. Kisselev AF, Callard A, Goldberg AL (2006) Importance of the different proteolytic sites of the proteasome and the efficacy of inhibitors varies with the protein substrate. J Biol Chem 281:8582–8590CrossRefPubMedGoogle Scholar
  32. Kwon YD, Nagy I, Adams PD, Baumeister W, Jap BK (2004) Crystal structures of the Rhodococcus proteasome with and without its pro-peptides: implications for the role of the pro-peptide in proteasome assembly. J Mol Biol 335:233–245CrossRefPubMedGoogle Scholar
  33. Levy F, Burri L, Morel S, Peitrequin AL, Levy N, Bachi A, Hellman U, Van den Eynde BJ, Servis C (2002) The final N-terminal trimming of a subaminoterminal proline-containing HLA class I-restricted antigenic peptide in the cytosol is mediated by two peptidases. J Immunol 169:4161–4171PubMedGoogle Scholar
  34. Liu C-W, Strickland E, DeMartino GN, Thomas PJ (2005) Recognition and processing of misfolded proteins by PA700, the 19S regulatory complex of the 26S proteasome. Methods Mol Biol:71–81Google Scholar
  35. Löwe J, Stock D, Jap B, Zwickl P, Baumeister W, Huber R (1995) Crystal structure of the 20 s proteasome from the archaeon T. acidophilum at 3.4 angstrom resolution. Science 268:533–539CrossRefPubMedGoogle Scholar
  36. Macpherson E, Tomkinson B, Balow RM, Hoglund S, Zetterqvist O (1987) Supramolecular structure of tripeptidyl peptidase II from human erythrocytes as studied by electron microscopy, and its correlation to enzyme activity. Biochem J 248:259–263PubMedGoogle Scholar
  37. Marcilla M, Cragnolini JJ, Lopez de Castro JAL (2007) Proteasome-independent HLA-B27 ligands arise mainly from small basic proteins. Mol Cell Proteomics 6:923–938CrossRefPubMedGoogle Scholar
  38. McKay RM, McKay JP, Suh JM, Avery L, Graff JM (2007) Tripeptidyl peptidase II promotes fat formation in a conserved fashion. EMBO Rep 8:1183–1189CrossRefPubMedGoogle Scholar
  39. Nakamura Y, Nakano K, Umehara T, Kimura M, Hayashizaki Y, Tanaka A, Horikoshi M, Padmanabhan B, Yokoyama S (2007) Structure of the oncoprotein gankyrin in complex with S6 ATPase of the 26S proteasome. Structure 15:179–189CrossRefPubMedGoogle Scholar
  40. Nickell S, Beck F, Korinek A, Mihalache O, Baumeister W, Plitzko JM (2007a) Automated cryoelectron microscopy of “single particles” applied to the 26S proteasome. FEBS Lett 581:2751–2756CrossRefPubMedGoogle Scholar
  41. Nickell S, Mihalache O, Beck F, Hegerl R, Korinek A, Baumeister W (2007b) Structural analysis of the 26S proteasome by cryoelectron tomography. Biochem Biophys Res Commun 353:115–120CrossRefPubMedGoogle Scholar
  42. Niedermann G (2002) Immunological functions of the proteasome. Curr Top Microbiol Immunol 268:91–136PubMedGoogle Scholar
  43. Ortega J, Heymann JB, Kajava AV, Ustrell V, Rechsteiner M, Steven AC (2005) The axial channel of the 20S proteasome opens upon binding of the PA200 activator. J Mol Biol 346:1221–1227CrossRefPubMedGoogle Scholar
  44. Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, Ferrin TE (2004) UCSF Chimera—a visualization system for exploratory research and analysis. J Comput Chem 25:1605–1612CrossRefPubMedGoogle Scholar
  45. Pickart CM, Cohen RE (2004) Proteasomes and their kin: proteases in the machine age. Nat Rev Mol Cell Biol 5:177–187CrossRefPubMedGoogle Scholar
  46. Pickart CM, VanDemark AP (2000) Opening doors into the proteasome. Nat Struct Biol 7:999–1001CrossRefPubMedGoogle Scholar
  47. Princiotta MF, Schubert U, Chen WS, Bennink JR, Myung J, Crews CM, Yewdell JW (2001) Cells adapted to the proteasome inhibitor 4-hydroxy5-iodo-3-nitrophenylacetyl-Leu-Leu-leucinal-vinyl sulfone require enzymatically active proteasomes for continued survival. Proc Natl Acad Sci U S A 98:513–518CrossRefPubMedGoogle Scholar
  48. Rechsteiner M, Realini C, Ustrell V (2000) The proteasome activator 11 S REG (PA28) and class I antigen presentation. Biochem J 345:1–15CrossRefPubMedGoogle Scholar
  49. Reits E, Neijssen J, Herberts C, Benckhuijsen W, Janssen L, Drijfhout JW, Neefjes J (2004) A major role for TPPII in trimming proteasomal degradation products for MHC class I antigen presentation. Immunity 20:495–506CrossRefPubMedGoogle Scholar
  50. Robinson CV, Sali A, Baumeister W (2007) The molecular sociology of the cell. Nature 450:973–982CrossRefPubMedGoogle Scholar
  51. Rockel B, Peters J, Kühlmorgen B, Glaeser RM, Baumeister W (2002) A giant protease with a twist: the TPP II complex from Drosophila studied by electron microscopy. EMBO J 21:5979–5984CrossRefPubMedGoogle Scholar
  52. Rockel B, Peters J, Müller SA, Seyit G, Ringler P, Hegerl R, Glaeser RM, Baumeister W (2005) Molecular architecture and assembly mechanism of Drosophila tripeptidyl peptidase II. Proc Natl Acad Sci U S A 102:10135–10140CrossRefPubMedGoogle Scholar
  53. Rose C, Vargas F, Facchinetti P, Bourgeat P, Bambal RB, Bishop PB, Chan SM, Moore AN, Ganellin CR, Schwartz JC (1996) Characterization and inhibition of a cholecystokinin-inactivating serine peptidase. Nature 380:403–409CrossRefPubMedGoogle Scholar
  54. Sanches M, Alves BSC, Zanchin NIT, Guimaraes BG (2007) The crystal structure of the human Mov34 MPN domain reveals a metal-free dimer. J Mol Biol 370:846–855CrossRefPubMedGoogle Scholar
  55. Schreiner P, Chen X, Husnjak K, Randles L, Zhang NX, Elsasser S, Finley D, Dikic I, Walters KJ, Groll M (2008) Ubiquitin docking at the proteasome through a novel pleckstrin-homology domain interaction. Nature 453:548–552CrossRefPubMedGoogle Scholar
  56. Seemüller E, Lupas A, Zühl F, Zwickl P, Baumeister W (1995a) The proteasome from Thermoplasma acidophilum is neither a cysteine nor a serine-protease. FEBS Lett 359:173–178CrossRefPubMedGoogle Scholar
  57. Seemüller E, Lupas A, Stock D, Löwe J, Huber R, Baumeister W (1995b) Proteasome from Thermoplasma acidophilum: a threonine protease. Science 268:579–583CrossRefPubMedGoogle Scholar
  58. Seifert U, Maranon C, Shmueli A, Desoutter JF, Wesoloski L, Janek K, Henklein P, Diescher S, Andrieu M, de la Salle H, Weinschenk T, Schild H, Laderach D, Galy A, Haas G, Kloetzel PM, Reiss Y, Hosmalin A (2003) An essential role for tripeptidyl peptidase in the generation of an MHC class I epitope. Nat Immunol 4:375–379CrossRefPubMedGoogle Scholar
  59. Seyit G, Rockel B, Baumeister W, Peters J (2006) Size matters for the tripeptidylpeptidase II complex from Drosophila—the 6-MDa spindle form stabilizes the activated state. J Biol Chem 281:25723–25733CrossRefPubMedGoogle Scholar
  60. Sharon M, Taverner T, Ambroggio XI, Deshaies RJ, Robinson CV (2006a) Structural organization of the 19S proteasome lid: Insights from MS of intact complexes. Plos Biol 4:1314–1323CrossRefGoogle Scholar
  61. Sharon M, Witt S, Felderer K, Rockel B, Baumeister W, Robinson CV (2006b) 20S proteasomes have the potential to keep substrates in store for continual degradation. J Biol Chem 281:9569–9575CrossRefPubMedGoogle Scholar
  62. Smith DM, Chang SC, Park S, Finley D, Cheng Y, Goldberg AL (2007) Docking of the proteasomal ATPases' carboxyl termini in the 20S proteasome's alpha ring opens the gate for substrate entry. Mol Cell 27:731–744CrossRefPubMedGoogle Scholar
  63. Stavropoulou V, Xie JJ, Henriksson M, Tomkinson B, Imreh S, Masucci MG (2005) Mitotic infidelity and centrosome duplication errors in cells overexpressing tripeptidyl-peptidase II. Cancer Res 65:1361–1368CrossRefPubMedGoogle Scholar
  64. Stavropoulou V, Vasquez V, Cereser B, Freda E, Masucci MG (2006) TPPII promotes genetic instability by allowing the escape from apoptosis of cells with activated mitotic checkpoints. Biochem Biophys Res Commun 346:415–425CrossRefPubMedGoogle Scholar
  65. Tamura T, Tamura N, Cejka Z, Hegerl R, Lottspeich F, Baumeister W (1996) Tricorn protease—the core of a modular proteolytic system. Science 274:1385–1389CrossRefPubMedGoogle Scholar
  66. Tamura N, Lottspeich F, Baumeister W, Tamura T (1998) The role of tricorn protease and its aminopeptidase-interacting factors in cellular protein degradation. Cell 95:637–648CrossRefPubMedGoogle Scholar
  67. Tomkinson B (1999) Tripeptidyl peptidases: enzymes that count. Trends Biochem Sci 24:355–359CrossRefPubMedGoogle Scholar
  68. Tomkinson B (2000) Association and dissociation of the tripeptidyl-peptidase II complex as a way of regulating the enzyme activity. Arch Biochem Biophys 376:275–280CrossRefPubMedGoogle Scholar
  69. Tomkinson B, Laoi BN, Wellington K (2002) The insert within the catalytic domain of tripeptidyl-peptidase II is important for the formation of the active complex. Eur J Biochem 269:1438–1443CrossRefPubMedGoogle Scholar
  70. Unno M, Mizushima T, Morimoto Y, Tomisugi Y, Tanaka K, Yasuoka N, Tsukihara T (2002) The structure of the mammalian 20S proteasome at 2.75 angstrom resolution. Structure 10:609–618CrossRefPubMedGoogle Scholar
  71. van Endert P (2008) Role of tripeptidyl peptidase II in MHC class I antigen processing—the end of controversies? Eur J Immunol 38:609–613CrossRefPubMedGoogle Scholar
  72. Voges D, Zwickl P, Baumeister W (1999) The 26S proteasome: a molecular machine designed for controlled proteolysis. Annu Rev Biochem 68:1015–1068CrossRefPubMedGoogle Scholar
  73. Walz J, Tamura T, Tamura N, Grimm R, Baumeister W, Koster AJ (1997) Tricorn protease exists as an icosahedral supermolecule in vivo. Mol Cell 1:59–65CrossRefPubMedGoogle Scholar
  74. Walz J, Erdmann A, Kania M, Typke D, Koster AJ, Baumeister W (1998) 26S proteasome structure revealed by three-dimensional electron microscopy. J Struct Biol 121:19–29CrossRefPubMedGoogle Scholar
  75. Walz J, Koster AJ, Tamura T, Baumeister W (1999) Capsids of tricorn protease studied by electron cryomicroscopy. J Struct Biol 128:65–68CrossRefPubMedGoogle Scholar
  76. Wang EW, Kessler BM, Borodovsky A, Cravatt BF, Bogyo M, Ploegh HL, Glas R (2000) Integration of the ubiquitin-proteasome pathway with a cytosolic oligopeptidase activity. Proc Natl Acad Sci U S A 97:9990–9995CrossRefPubMedGoogle Scholar
  77. Wang Q, Young P, Walters KJ (2005) Structure of S5a bound to monoubiquitin provides a model for polyubiquitin recognition. J Mol Biol 348:727–739CrossRefPubMedGoogle Scholar
  78. Whitby FG, Masters EI, Kramer L, Knowlton JR, Yao Y, Wang CC, Hill CP (2000) Structural basis for the activation of 20S proteasomes by 11S regulators. Nature 408:115–120CrossRefPubMedGoogle Scholar
  79. Yao TT, Cohen RE (1999) Giant proteases: beyond the proteasome. Curr Biol 9:R551–R553CrossRefPubMedGoogle Scholar
  80. York IA, Bhutani N, Zendzian S, Goldberg AL, Rock KL (2006) Tripeptidyl peptidase II is the major peptidase needed to trim long antigenic precursors, but is not required for most MHC class I antigen presentation. J Immunol 177:1434–1443PubMedGoogle Scholar
  81. Yoshimura T, Kameyama K, Takagi T, Ikai A, Tokunaga F, Koide T, Tanahashi N, Tamura T, Cejka Z, Baumeister W, Tanaka K, Ichihara A (1993) Molecular characterization of the 26S proteasome complex from rat-liver. J Struct Biol 111:200–211CrossRefPubMedGoogle Scholar
  82. Zwickl P, Seemüller E, Kapelari B, Baumeister W (2001) The proteasome: a supramolecular assembly designed for controlled proteolysis. Adv Protein Chem 59:187–222CrossRefPubMedGoogle Scholar

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© Springer-Verlag 2008

Authors and Affiliations

  1. 1.Department of Molecular Structural BiologyMax-Planck Institute of BiochemistryMartinsriedGermany

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