Skip to main content

The balance of ubiquitin conjugation and deconjugation

Objective

Ubiquitin conjugation is one of the most important signaling systems in the eukaryotic cell. Different types of mono- and polyubiquitin chains determine the fate of target proteins by redirecting them for degradation, relocalization or interaction with new partners. The type of ubiquitin modification on any target is determined by the interplay between the conjugating E2/E3 complexes on the one hand and deubiquitinating enzymes on the other. In practice, it is the balance between conjugating and deconjugating systems that determines the result of the various ubiquitination signals. For three different regulatory systems that are critical for correct genome maintenance, we are now in a position to study not just the individual process of conjugation or deconjugation in isolation, but rather, reconstitute the entire reaction on a defined physiological target. These three target systems, histone H2A, PCNA and P53, can be mono-ubiquitinated by a defined E3-ligase, poly-ubiquitinated by a second ligase and deconjugated by defined deubiquitinating enzymes in a reaction that is affected by known allosteric modulators. Our unique collection of tools to study these systems in vitro allows reconstitution of the full reaction, to trap intermediates, and to study their interaction from atomic detail to kinetic reactivity. Using X-ray crystallography of critical intermediates and kinetic analysis of individual reactions by FRET and surface plasmon resonance, we can address how the mono-, poly and deubiquitinating reactions affect each other. By answering mechanistic questions on the relative effect of the forward and backward reaction components and their modulators we will provide a solid basis for drug design studies that target these pathways against cancer development.

Field of science

  • /natural sciences/earth and related environmental sciences/geology/mineralogy/crystallography
  • /natural sciences/biological sciences/biochemistry/biomolecules/proteins
  • /medical and health sciences/clinical medicine/oncology/cancer
  • /natural sciences/biological sciences/genetics and heredity/genome
  • /natural sciences/biological sciences/biochemistry/biomolecules/proteins/enzymes

Call for proposal

ERC-2009-AdG
See other projects for this call

Funding Scheme

ERC-AG - ERC Advanced Grant

Host institution

STICHTING HET NEDERLANDS KANKER INSTITUUT-ANTONI VAN LEEUWENHOEK ZIEKENHUIS
Address
Plesmanlaan 121
1066 CX Amsterdam
Netherlands
Activity type
Research Organisations
EU contribution
€ 2 299 720
Principal investigator
Titia Karen Sixma (Prof.)
Administrative Contact
Henri Van Luenen (Dr.)

Beneficiaries (1)

STICHTING HET NEDERLANDS KANKER INSTITUUT-ANTONI VAN LEEUWENHOEK ZIEKENHUIS
Netherlands
EU contribution
€ 2 299 720
Address
Plesmanlaan 121
1066 CX Amsterdam
Activity type
Research Organisations
Principal investigator
Titia Karen Sixma (Prof.)
Administrative Contact
Henri Van Luenen (Dr.)