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MRC OIRO has access to a state-of-the-art proteomics platform at the Target Discovery Institute (TDI) / Centre for Medicines Discovery (CMD) within the Nuffield Department of Medicine (NDM). The Kessler group has previously collaborated with MRC/Oncology PIs to unravel molecular insights underpinning cellular alterations caused by IR and DNA damaging agents, resulting in >40 joint publications and is a valued collaborator in using this core.

Figure 1: Discovery proteomics as part of the Kessler group in the TDI/CMD will provide the Proteomics Core B for the MRC/UKRI OIRO Unit. This reflects substantial added value to OIRO’s research programmes

Discovery Proteomics Facility DPF: The Discovery Proteomics Facility (DPF) is a non-core funded small research facility (SRF) consisting of six members of staff who are in charge of operating three orbitrap mass spectrometers as well as a timsTOF instrument. The DPF is located of the TDI, Centre for Medicines Discovery CMD, NDMRB Building, within 1 minute walking distance from the Old Road Campus Research Building ORCRB with the OIRO MRC Unit. The DPF SRF team, led by Prof Roman Fischer (https://www.tdi.ox.ac.uk/team/roman-fischer) and Dr Iolanda Vendrell, is in constant interaction with the Kessler group, which helps driving new innovation through biology driven needs for improving MS workflows. The DPF has established a panel of MS methods (e.g., deep proteomics/prefractionation, PTM analysis, PRM methodology, phosphoproteomics, ubiquitomics, and acetylomics) and can offer training to MRC/OIRO researchers. The DPF has available high-end mass spectrometry instrumentation (Orbitrap Fusion Lumos, QE-HF, QE-classic and a TimsTOF Pro) dedicated to proteomics. Workflows to improve PTM detection and localisation, small molecule compound profiling by chemoproteomics or detectable proteome depth are under constant development to tailor proteomics experiments, in particular for the discovery of molecular changes in cells exposed to ionising radiation.

Past and Future Research using Proteomics technology: Benedikt Kessler’s group is focused on ubiquitin and protease biology and mass spectrometry, proteomics, reflecting considerable synergy with MRC’s research on ubiquitin mediated processes related to IR and replication stress (Ramadan, D’Angiolella and Hammond). The Kessler Lab has developed chemoproteomics methods to profile active ubiquitin processing enzymes such as deubiquitylating enzymes (DUBs) and the dynamic ubiquitome for the characterisation of USP7 inhibitors as novel potential therapeutic agents in multiple myeloma (1) and how USP18 modulates the interferon (IFN) response and convers sensitivity to ionising radiation in cancer cells (2). In particular, key contributions were made to understand the role of SPRTN protease in resolving DNA-protein crosslinks in IR DDR with the Ramadan group (1) and cyclin F ubiquitin E3 ligase in DNA replication stress with the D’Angiolella team (3).

This core has previously established an anti-GlyGly based enrichment strategy of ubiquitinated proteins by mass spectrometry (2,4,5). In addition, it has recently implemented the UbiFast ubiquitomics approach that reduces the need of input biological material (6), and can combine this with a data independent analysis (DIA) method for enhanced detection of ubiquitylation sites (7). These advances in MS methodologies, in particular including DIA, will also be beneficial for peptide mapping of intracellular proteins (Olcina, Giaccia PLs) or AP-MS based interactomics (Moon & Humphrey PLs). Dr Iolanda Vendrell will directly collaborate with group members of the PLs to implement these MS workflows in the respective research programmes.

References:

1. Turnbull A Nature 2017 Oct 26; 550 (7677):481-486.;

2 . Pinto-Fernandez A BJC 2021 Feb; 124(4):817-83; (

3 . Vaz B Mol Cell 2016 Nov 17; 64(4):704-719;

4 . Burdova K EMBO J 2019 Oct 15; 38(20):e101443;

5 . Sapmaz A et al., Nat Commun. 2019 Mar 29;10(1):1454.;

6 . Unsworth AJ et al., Thromb Haemost. 2019 Jan;119(1):104-116.;

7 . Udeshi ND et al., Nat Commun. 2020 Jan 17;11(1):359.;

8 . Steger M et al., bioRxiv https://doi.org/ 10.1101/2020.07.23.218651