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dc.creatorVizovisek, Matej
dc.creatorVidmar, Robert
dc.creatorVan, Quickelberghe Emmy
dc.creatorImpens, Francis
dc.creatorAnđelković, Uroš
dc.creatorSobotic, Barbara
dc.creatorStoka, Veronika
dc.creatorGevaert, Kris
dc.creatorTurk, Boris
dc.creatorFonovic, Marko
dc.date.accessioned2019-01-30T17:43:37Z
dc.date.available2019-01-30T17:43:37Z
dc.date.issued2015
dc.identifier.issn1615-9853
dc.identifier.urihttp://cer.ihtm.bg.ac.rs/handle/123456789/1626
dc.description.abstractProteases are important effectors of numerous physiological and pathological processes. Reliable determination of a protease's specificity is crucial to understand protease function and to develop activity-based probes and inhibitors. During the last decade, various proteomic approaches for profiling protease substrate specificities were reported. Although most of these approaches can identify up to thousands of substrate cleavage events in a single experiment, they are often time consuming and methodologically challenging as some of these approaches require rather complex sample preparation procedures. For such reasons their application is often limited to those labs that initially introduced them. Here, we report on a fast and simple approach for proteomic profiling of protease specificities (fast profiling of protease specificity (FPPS)), which can be applied to complex protein mixtures. FPPS is based on trideutero-acetylation of novel N-termini generated by the action of proteases and subsequent peptide fractionation on Stage Tips containing ion-exchange and reverse phase chromatographic resins. FPPS can be performed in 2 days and does not require extensive fractionation steps. Using this approach, we have determined the specificity profiles of the cysteine cathepsins K, L and S. We further validated our method by comparing the results with the specificity profiles obtained by the N-terminal combined fractional diagonal chromatography method. This comparison pointed to almost identical substrate specificities for all three cathepsins and confirmed the reliability of the FPPS approach. All MS data have been deposited in the ProteomeXchange with identifiers PXD001536 and PXD001553 (; ).en
dc.publisherWiley-Blackwell, Hoboken
dc.relationSlovenian Research Agency - P1-0140
dc.relationSlovenian Research Agency - J1-3602
dc.relationSlovenian Research Agency - N1-0022
dc.relationSlovenian Research Agency - J1-0185
dc.relationResearch Foundation - Flanders (FWO-Vlaanderen) - G.0048.08
dc.relationResearch Foundation - Flanders (FWO-Vlaanderen) - G.0C37.14N
dc.rightsrestrictedAccess
dc.sourceProteomics
dc.subjectCathepsin protease specificityen
dc.subjectIntact protein-based cleavage site discoveryen
dc.subjectN-terminomicsen
dc.subjectTechnologyen
dc.titleFast profiling of protease specificity reveals similar substrate specificities for cathepsins K, L and Sen
dc.typearticle
dc.rights.licenseARR
dcterms.abstractВизовисек, Матеј; Геваерт, Крис; Турк, Борис; Фоновиц, Марко; Ван, Qуицкелбергхе Еммy; Импенс, Францис; Соботиц, Барбара; Стока, Вероника; Aнђелковић, Урош; Видмар, Роберт;
dc.citation.volume15
dc.citation.issue14
dc.citation.spage2479
dc.citation.epage2490
dc.citation.other15(14): 2479-2490
dc.citation.rankM21
dc.description.otherAccepted version: [http://cer.ihtm.bg.ac.rs/handle/123456789/3209]
dc.identifier.pmid25626674
dc.identifier.doi10.1002/pmic.201400460
dc.identifier.rcubConv_3369
dc.identifier.scopus2-s2.0-84924390096
dc.identifier.wos000357946600011
dc.type.versionpublishedVersion


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