Objective This research proposal describes an ambitious effort to characterize structurally and biochemically ubiquitin chain initiation and elongation by the anaphase-promoting complex or cyclosome (APC/C) in complex with two well-characterized substrates. The APC/C is a multi-subunit cullin-RING E3 ubiquitin ligase that controls progression through the cell cycle by a temporal regulation of its activity and substrate specificity. Regulation and specificity of this E3 ligase is achieved through mutually exclusive binding of two structurally related co-activator subunits termed Cdc20 and Cdh1, as well as through APC/C inhibitors, varying substrate affinities and auto-ubiquitylation of its cognate E2s, namely UbcH10 and Ube2S. In order to understand ubiquitin chain initiation and elongation of the two well-known APC/C substrates cyclin B and securin, I am aiming to use a combined approach of cryo-electron microscopy, X-ray crystallography and a variety of biochemical methods. Within this project I will use cryo-electron microscopy studies to uncover the molecular mechanisms of substrate recognition and ubiquitin chain initiation and elongation by analyzing the APC/CCdh1 co-activator complex bound to its transiently associated E2 enzymes Ube2S or UbcH10 and one of the aforementioned high affinity substrates. Crystallization of selected sub-complexes, namely the catalytic core of the APC/C (composed of Apc2 and Apc11) is intended. If obtained, this high-resolution information will then assist the interpretation of the resulting density maps derived from cryo-electron microscopy. Fields of science medical and health sciencesbasic medicinemedicinal chemistrynatural sciencesearth and related environmental sciencesgeologymineralogycrystallographynatural sciencesphysical sciencesopticsmicroscopynatural sciencesbiological sciencesbiochemistrybiomoleculesproteinsenzymesnatural sciencescomputer and information sciencesdata sciencedata processing Programme(s) H2020-EU.1.3. - EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions Main Programme H2020-EU.1.3.2. - Nurturing excellence by means of cross-border and cross-sector mobility Topic(s) MSCA-IF-2014-EF - Marie Skłodowska-Curie Individual Fellowships (IF-EF) Call for proposal H2020-MSCA-IF-2014 See other projects for this call Funding Scheme MSCA-IF-EF-ST - Standard EF Coordinator UNITED KINGDOM RESEARCH AND INNOVATION Net EU contribution € 183 454,80 Address POLARIS HOUSE NORTH STAR AVENUE SN2 1FL Swindon United Kingdom See on map Region South West (England) Gloucestershire, Wiltshire and Bristol/Bath area Swindon Activity type Research Organisations Links Contact the organisation Opens in new window Website Opens in new window Participation in EU R&I programmes Opens in new window HORIZON collaboration network Opens in new window Total cost € 183 454,80
