Skip to main content
SpringerLink
Log in

Thioester and Oxyester Linkages in the Ubiquitin System

  • Protocol
  • First Online:
The Ubiquitin Code

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2602))

Abstract

The traditional textbook describes ubiquitylation as the conjugation of ubiquitin to a target by forming a covalent bond connecting ubiquitin’s carboxy-terminal glycine residue with an acceptor amino acid like lysine or amino-terminal methionine in the substrate protein. While this adequately depicts a significant fraction of cellular ubiquitylation processes, a growing number of ubiquitin modifications do not follow this rule. Recent data demonstrate that ubiquitin can also be efficiently attached to other amino acids, such as cysteine, serine, and threonine, via ester bonding. Initially observed for a virus-encoded ubiquitin ligase, which targets a cysteine residue in a host protein to initiate its degradation, ester-linked ubiquitylation is now shown to also drive regular cellular processes. These ubiquitylation events expand the complexity and diversity of ubiquitin signaling and broaden the capability of cellular messages in the so-called ubiquitin code. Still, questions on the prevalence, relevance, and involvement in physiological and cellular functions await clearing. In this review, we aim to summarize our knowledge on ester-linked ubiquitylation and introduce experimental strategies to circumvent technical issues that complicate analysis of this uncommon posttranslational modification.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 199.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Scheffner M, Nuber U, Huibregtse JM (1995) Protein ubiquitination involving an E1–E2–E3 enzyme ubiquitin thioester cascade. Nature 373:81–83

    Article  CAS  PubMed  Google Scholar 

  2. Chau V, Tobias JW, Bachmair A et al (1989) A Multiubiquitin Chain Is Confined to Specific Lysine in a Targeted Short-Lived Protein. Science 80(243):1576–1583

    Article  Google Scholar 

  3. Xu P, Duong DM, Seyfried NT et al (2009) Quantitative proteomics reveals the function of unconventional ubiquitin chains in proteasomal degradation. Cell 137:133–145

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Akutsu M, Dikic I, Bremm A (2016) Ubiquitin chain diversity at a glance. J Cell Sci 129:875–880

    CAS  PubMed  Google Scholar 

  5. Komander D, Rape M (2012) The ubiquitin code. Annu Rev Biochem 81:203–229

    Article  CAS  PubMed  Google Scholar 

  6. Cadwell K, Coscoy L (2005) Ubiquitination on nonlysine residues by a viral E3 ubiquitin ligase. Science 80(309):127–130

    Article  Google Scholar 

  7. Carvalho AF, Pinto MP, Grou CP et al (2007) Ubiquitination of mammalian Pex5p, the peroxisomal import receptor. J Biol Chem 282:31267–31272

    Article  CAS  PubMed  Google Scholar 

  8. Williams C, Van Den BM, Sprenger RR et al (2007) A conserved cysteine is essential for Pex4p-dependent ubiquitination of the peroxisomal import receptor Pex5p. J Biol Chem 282:22534–22543

    Article  CAS  PubMed  Google Scholar 

  9. Vosper JMD, McDowell GS, Hindley CJ et al (2009) Ubiquitylation on canonical and non-canonical sites targets the transciption factor neurogenin for ubiquitin-mediated proteolysis. J Biol Chem 284:15458–15468

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Wang X, Herr RA, Hansen TH (2012) Ubiquitination of substrates by esterification. Traffic 13:19–24

    Article  CAS  PubMed  Google Scholar 

  11. McDowell GS, Philpott A (2013) Non-canonical ubiquitylation: Mechanisms and consequences. Int J Biochem Cell Biol 45:1833–1842

    Article  CAS  PubMed  Google Scholar 

  12. McClellan AJ, Laugesen SH, Ellgaard L (2019) Cellular functions and molecular mechanisms of non-lysine ubiquitination. Open Biol 9:190147

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Kalayil S, Bhogaraju S, Bonn F et al (2018) Insights into catalysis and function of phosphoribosyl-linked serine ubiquitination. Nature 557:734–738

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Tokarev AA, Munguia J, Guatelli JC (2011) Serine-threonine ubiquitination mediates downregulation of BST-2/tetherin and relief of restricted virion release by HIV-1 Vpu. J Virol 85:51–63

    Article  CAS  PubMed  Google Scholar 

  15. Gu H, Fada BJ (2020) Specificity in ubiquitination triggered by virus infection. Int J Mol Sci 21(11):4088

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Breitschopf K, Bengal E, Ziv T et al (1998) A novel site for ubiquitination: the N-terminal residue, and not internal lysines of MyoD, is essential for conjugation and degradation of the protein. EMBO J 17:5964–5973

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Dittmar G, Winklhofer KF (2020) Linear ubiquitin chains: cellular functions and strategies for detection and quantification. Front Chem 7:915

    Article  PubMed  PubMed Central  Google Scholar 

  18. Peter DM, Vögeli B, Cortina NS et al (2016) A chemo-enzymatic road map to the synthesis of CoA esters. Molecules 21(4):517

    Article  PubMed  PubMed Central  Google Scholar 

  19. Plechanovova A, Jaffray EG, Tatham MH et al (2012) Structure of a RING E3 ligase and ubiquitin-loaded E2 primed for catalysis. Nature 489:115

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Ravid T, Hochstrasser M (2007) Autoregulation of an E2 enzyme by ubiquitin-chain assembly on its catalytic residue. Nat Cell Biol 94(9):422–427

    Article  Google Scholar 

  21. Song J, Wang J, Jozwiak AA et al (2009) Stability of thioester intermediates in ubiquitin-like modifications. Protein Sci 18:2492–2499

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. De Cesare V, Carbajo Lopez D, Mabbitt PD et al (2021) Deubiquitinating enzyme amino acid profiling reveals a class of ubiquitin esterases. Proc Natl Acad Sci U S A 118:e2006947118

    Google Scholar 

  23. Creasy DM, Cottrell JS (2004) Unimod: Protein modifications for mass spectrometry. Proteomics 4:1534–1536

    Article  CAS  PubMed  Google Scholar 

  24. Udeshi ND, Mertins P, Svinkina T et al (2013) Large-scale identification of ubiquitination sites by mass spectrometry. Nat Protoc 8:1950

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Udeshi ND, Svinkina T, Mertins P et al (2013) Refined preparation and use of anti-diglycine remnant (k-ε-gg) antibody enables routine quantification of 10,000s of ubiquitination sites in single proteomics experiments. Mol Cell Proteomics 12:825–831

    Article  CAS  PubMed  Google Scholar 

  26. Udeshi ND, Mani DC, Satpathy S et al (2020) Rapid and deep-scale ubiquitylation profiling for biology and translational research. Nat Commun 11(1):359

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Wagner SA, Beli P, Weinert BT et al (2012) Proteomic analyses reveal divergent ubiquitylation site patterns in murine tissues. Mol Cell Proteomics 11:1578–1585

    Article  PubMed  PubMed Central  Google Scholar 

  28. Akimov V, Barrio-Hernandez I, Hansen SVF et al (2018) UbiSite approach for comprehensive mapping of lysine and N-terminal ubiquitination sites. Nat Struct Mol Biol 25:631–640

    Article  CAS  PubMed  Google Scholar 

  29. Clague MJ, Urbé S, Komander D (2019) Breaking the chains: deubiquitylating enzyme specificity begets function. Nat Rev Mol Cell Biol 20:338–352

    Article  CAS  PubMed  Google Scholar 

  30. Jarosch E, Lenk U, Sommer T (2003) Endoplasmic reticulum-associated protein degradation. Int Rev Cytol 223:39–81

    Article  CAS  PubMed  Google Scholar 

  31. Shimizu Y, Okuda-Shimizu Y, Hendershot LM (2010) Ubiquitylation of an ERAD substrate occurs on multiple types of amino acids. Mol Cell 40:917–926

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Boban M, Ljungdahl PO, Foisner R (2015) Atypical ubiquitylation in yeast targets lysine-less Asi2 for proteasomal degradation. J Biol Chem 290:2489–2495

    Article  CAS  PubMed  Google Scholar 

  33. Weber A, Cohen I, Popp O et al (2016) Sequential poly-ubiquitylation by specialized conjugating enzymes expands the versatility of a quality control ubiquitin ligase. Mol Cell 63:827–839

    Article  CAS  PubMed  Google Scholar 

  34. Lips C, Ritterhoff T, Weber A et al (2020) Who with whom: functional coordination of E2 enzymes by RING E3 ligases during poly-ubiquitylation. EMBO J 39(22):e104863

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Wang X, Herr RA, Rabelink M et al (2009) Ube2j2 ubiquitinates hydroxylated amino acids on ER-associated degradation substrates. J Cell Biol 187:655–668

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Tan JME, Cook ECL, Van Den Berg M et al (2019) Differential use of E2 ubiquitin conjugating enzymes for regulated degradation of the rate-limiting enzymes HMGCR and SQLE in cholesterol biosynthesis. Atherosclerosis 281:137–142

    Article  CAS  PubMed  Google Scholar 

  37. Chua NK, Hart-Smith G, Brown AJ (2019) Non-canonical ubiquitination of the cholesterol-regulated degron of squalene monooxygenase. J Biol Chem 294:8134–8147

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Ishikura S, Weissman AM, Bonifacino JS (2010) Serine residues in the cytosolic tail of the T-cell antigen receptor α-chain mediate ubiquitination and endoplasmic reticulum-associated degradation of the unassembled protein. J Biol Chem 285:23916–23924

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Burr ML, Cano F, Svobodova S et al (2011) HRD1 and UBE2J1 target misfolded MHC class I heavy chains for endoplasmic reticulum-associated degradation. Proc Natl Acad Sci U S A 108:2034–2039

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Golnik R, Lehmann A, Kloetzel PM et al (2016) Major histocompatibility complex (MHC) class i processing of the NY-ESO-1 antigen is regulated by Rpn10 and Rpn13 Proteins and immunoproteasomes following non-lysine ubiquitination. J Biol Chem 291:8805–8815

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Yuan L, Hassan BA (2014) Neurogenins in brain development and disease: an overview. Arch Biochem Biophys 558:10–13

    Article  CAS  PubMed  Google Scholar 

  42. McDowell GS, Kucerova R, Philpott A (2010) Non-canonical ubiquitylation of the proneural protein Ngn2 occurs in both Xenopus embryos and mammalian cells. Biochem Biophys Res Commun 400:655–660

    Article  CAS  PubMed  Google Scholar 

  43. Roark R, Itzhaki L, Philpott A (2012) Complex regulation controls Neurogenin3 proteolysis. Biol Open 1:1264–1272

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Gilkerson J, Kelley DR, Tam R et al (2015) Lysine residues are not required for proteasome- mediated proteolysis of the auxin/indole acidic acid protein IAA1. Plant Physiol 168:708–720

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Peeler JC, Schedin-Weiss S, Soula M et al (2017) Isopeptide and ester bond ubiquitination both regulate degradation of the human dopamine receptor 4. J Biol Chem 292:21623–21630

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Skieterska K, Rondou P, Lintermans B et al (2015) KLHL12 Promotes Non-Lysine Ubiquitination of the dopamine receptors D4.2 and D4.4, but not of the ADHD-associated D4.7 variant. PLoS One 10(12):e0145654

    Article  PubMed  PubMed Central  Google Scholar 

  47. Kelsall IR, Zhang J, Knebel A et al (2019) The E3 ligase HOIL-1 catalyses ester bond formation between ubiquitin and components of the Myddosome in mammalian cells. Proc Natl Acad Sci U S A 116:13293–13298

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Petrova T, Zhang J, Nanda SK et al (2021) HOIL-1-catalysed, ester-linked ubiquitylation restricts IL-18 signaling in cytotoxic T cells but promotes TLR signalling in macrophages. FEBS J 288:5909–5924

    Article  CAS  PubMed  Google Scholar 

  49. Carvajal AR, Grishkovskaya I, Diaz CG et al (2021) The linear ubiquitin chain assembly complex (LUBAC) generates heterotypic ubiquitin chains. Elife 10:1–28

    Google Scholar 

  50. Platta HW, El MF, Bäumer BE et al (2009) Pex2 and Pex12 function as protein-ubiquitin ligases in peroxisomal protein import. Mol Cell Biol 29:5505–5516

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Okumoto K, Misono S, Miyata N et al (2011) Cysteine ubiquitination of PTS1 receptor Pex5p regulates Pex5p recycling. Traffic 12:1067–1083

    Article  CAS  PubMed  Google Scholar 

  52. Schwartzkopff B, Platta HW, Hasan S et al (2015) Cysteine-specific ubiquitination protects the peroxisomal import receptor Pex5p against proteasomal degradation. Biosci Rep 35:1–12

    Article  CAS  Google Scholar 

  53. Debelyy MO, Platta HW, Saffian D et al (2011) Ubp15p, a ubiquitin hydrolase associated with the peroxisomal export machinery. J Biol Chem 286:28223–28234

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Hensel A, Beck S, El MF et al (2011) Cysteine-dependent ubiquitination of Pex18p is linked to cargo translocation across the peroxisomal membrane. J Biol Chem 286:43495–43505

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. El MF, Brinkmeier R, Schrötter A et al (2013) Distinct ubiquitination cascades act on the peroxisomal targeting signal type 2 co-receptor pex18p. Traffic 14:1290–1301

    Article  Google Scholar 

  56. Tait SWG, De VE, Maas C et al (2007) Apoptosis induction by Bid requires unconventional ubiquitination and degradation of its N-terminal fragment. J Cell Biol 179:1453–1466

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Pao KC, Wood NT, Knebel A et al (2018) Activity-based E3 ligase profiling uncovers an E3 ligase with esterification activity. Nature 556:381–385

    Article  CAS  PubMed  Google Scholar 

  58. Mabbitt PD, Loreto A, Déry MA et al (2020) Structural basis for RING-Cys-Relay E3 ligase activity and its role in axon integrity. Nat Chem Biol 16:1227–1236

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Max-Delbrück-Center for Molecular Medicine in the Helmholz Association, Berlin Buch, Germany

    Alba Ferri-Blazquez, Ernst Jarosch & Thomas Sommer

  2. Institute for Biology, Humboldt-Universität zu Berlin, Berlin, Germany

    Alba Ferri-Blazquez & Thomas Sommer

Authors
  1. Alba Ferri-Blazquez
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ernst Jarosch
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Thomas Sommer
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Alba Ferri-Blazquez .

Editor information

Editors and Affiliations

  1. LCC-UPR 8241-CNRS Equipe M, Laboratoire de Chimie de Coordination, Toulouse, France

    Manuel S Rodriguez

  2. CIC bioGUNE, Derio, Spain

    Rosa Barrio

Rights and permissions

Reprints and permissions

Copyright information

© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Ferri-Blazquez, A., Jarosch, E., Sommer, T. (2023). Thioester and Oxyester Linkages in the Ubiquitin System. In: Rodriguez, M.S., Barrio, R. (eds) The Ubiquitin Code. Methods in Molecular Biology, vol 2602. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2859-1_1

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-2859-1_1

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2858-4

  • Online ISBN: 978-1-0716-2859-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation