Detailed knowledge of the molecular mechanisms underlying ubiquitin processing provided an inroad for designing molecular

probes targeting conjugating and deconjugating enzymes. This approach has regained interest also because it allows small molecule inhibitor development within the UPS [42]. Whereas ubiquitin processing enzymes (USPs, UCHs, OTUs) can be readily profiled using ubiquitin based chemical probes targeting proteolytic catalysis, the application of this activity-based approach towards the ubiquitin conjugating GSK2118436 cascade has proven to be challenging [43 and 44], although chemical crosslinking was successfully used for HECT domain E3 ligases [45]. In the case of deubiquitination, further progress has been made to create molecular probes mimicking different isopeptide linkages between ubiquitin and protein substrates including ubiquitin itself by integrating peptides at the P′ side of the scissile bond with an electrophilic moiety in the center, which appear to selectively target subsets of DUBs in crude cell extracts [46]. This approach will complement predominantly in vitro studies on how DUBs can distinguish between different poly-ubiquitin chains for processing, currently achieved using model substrates [47] or fluorescence resonance techniques [48]. Also, more recent

probes on the basis of the ubiquitin scaffold are now available with BKM120 ic50 C-terminal PLEK2 fluorescent moieties via ‘Click Chemistry’ [49], or N-terminal fluorescent or photoreactive moieties through total synthesis [50 and 51•]. Such tools will undoubtedly be used to profile ubiquitin processing enzymes such as DUBs, but also related enzymes specifically recognizing ubiquitin-like proteins, and thereby contribute to our understanding the role of these enzymes within the ubiquitin network in normal physiology as well as disease pathogenesis (Figure 4). The author declares no conflict of interest

in relation to the work described in this manuscript. Papers of particular interest, published within the period of review, have been highlighted as: • of special interest I am grateful for all the helpful discussions with colleagues and would like to apologise for all references that were not cited because of space constrains. “
“Current Opinion in Chemical Biology 2013, 17:73–82 This review comes from a themed issue on Omics Edited by Matthew Bogyo and Pauline M Rudd For a complete overview see the Issue and the Editorial Available online 6th January 2013 1367-5931/$ – see front matter, © 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.cbpa.2012.11.025 Proteomics has made astonishing advances in all areas including peptide enrichment, fractionation, mass spectrometry and data analysis — many of which are reviewed in this issue.