{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,25]],"date-time":"2026-03-25T07:20:05Z","timestamp":1774423205012,"version":"3.50.1"},"reference-count":41,"publisher":"Wiley","issue":"21","license":[{"start":{"date-parts":[[2023,8,29]],"date-time":"2023-08-29T00:00:00Z","timestamp":1693267200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["chemistry-europe.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["ChemBioChem"],"published-print":{"date-parts":[[2023,11,2]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Activity\u2010based protein profiling is a powerful chemoproteomic technique to detect active enzymes and identify targets and off\u2010targets of drugs. Here, we report the use of carmofur\u2010 and activity\u2010based probes to identify biologically relevant enzymes in the bacterial pathogen <jats:italic>Staphylococcus aureus<\/jats:italic>. Carmofur is an anti\u2010neoplastic prodrug of 5\u2010fluorouracil and also has antimicrobial and anti\u2010biofilm activity. Carmofur probes were originally designed to target human acid ceramidase, a member of the NTN hydrolase family with an active\u2010site cysteine nucleophile. Here, we first profiled the targets of a fluorescent carmofur probe in live <jats:italic>S. aureus<\/jats:italic> under biofilm\u2010promoting conditions and in liquid culture, before proceeding to target identification by liquid chromatography\/mass spectrometry. Treatment with a carmofur\u2010biotin probe led to enrichment of 20 enzymes from diverse families awaiting further characterization, including the NTN hydrolase\u2010related IMP cyclohydrolase PurH. However, the probe preferentially labeled serine hydrolases, thus displaying a reactivity profile similar to that of carbamates. Our results suggest that the electrophilic <jats:italic>N<\/jats:italic>\u2010carbamoyl\u20105\u2010fluorouracil scaffold could potentially be optimized to achieve selectivity towards diverse enzyme families. The observed promiscuous reactivity profile suggests that the clinical use of carmofur presumably leads to inactivation of a number human and microbial enzymes, which could lead to side effects and\/or contribute to therapeutic efficacy.<\/jats:p>","DOI":"10.1002\/cbic.202300473","type":"journal-article","created":{"date-parts":[[2023,8,8]],"date-time":"2023-08-08T09:13:21Z","timestamp":1691486001000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":10,"title":["Activity\u2010Based Protein Profiling in Methicillin\u2010Resistant <i>Staphylococcus aureus<\/i> Reveals the Broad Reactivity of a Carmofur\u2010Derived Probe"],"prefix":"10.1002","volume":"24","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3349-7741","authenticated-orcid":false,"given":"Md Jalal","family":"Uddin","sequence":"first","affiliation":[{"name":"Department of Medical Biology UiT- The Arctic University of Norway  9019 Troms\u00f8 Norway"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hermen S.","family":"Overkleeft","sequence":"additional","affiliation":[{"name":"Department of Bioorganic Synthesis Leiden Institute of Chemistry Leiden University  Einsteinweg 55 2333 CC Leiden The Netherlands"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7284-2264","authenticated-orcid":false,"given":"Christian S.","family":"Lentz","sequence":"additional","affiliation":[{"name":"Department of Medical Biology UiT- The Arctic University of Norway  9019 Troms\u00f8 Norway"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2023,8,29]]},"reference":[{"key":"e_1_2_9_1_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.aaf7497"},{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-4939-6439-0_1"},{"key":"e_1_2_9_3_1","first-page":"253","volume-title":"Recent Advances in Activity-Based Protein Profiling of Proteases","author":"Chakrabarty S.","year":"2019"},{"key":"e_1_2_9_4_1","doi-asserted-by":"publisher","DOI":"10.1515\/hsz-2019-0262"},{"key":"e_1_2_9_5_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cbpa.2020.06.011"},{"key":"e_1_2_9_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.chembiol.2015.12.003"},{"key":"e_1_2_9_7_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.96.26.14694"},{"key":"e_1_2_9_8_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41589-018-0060-1"},{"key":"e_1_2_9_9_1","doi-asserted-by":"publisher","DOI":"10.1039\/C5CC00356C"},{"key":"e_1_2_9_10_1","doi-asserted-by":"publisher","DOI":"10.1021\/jm301879g"},{"key":"e_1_2_9_11_1","doi-asserted-by":"publisher","DOI":"10.18632\/oncotarget.22637"},{"key":"e_1_2_9_12_1","doi-asserted-by":"publisher","DOI":"10.1021\/acs.chemrev.6b00209"},{"key":"e_1_2_9_13_1","doi-asserted-by":"publisher","DOI":"10.1002\/ddr.21984"},{"key":"e_1_2_9_14_1","doi-asserted-by":"publisher","DOI":"10.3748\/wjg.v20.i42.15682"},{"key":"e_1_2_9_15_1","doi-asserted-by":"publisher","DOI":"10.1038\/nrc1074"},{"key":"e_1_2_9_16_1","first-page":"1613","volume":"26","author":"Sato S.","year":"1999","journal-title":"Gan to Kagaku Ryoho"},{"key":"e_1_2_9_17_1","doi-asserted-by":"publisher","DOI":"10.1038\/srep01035"},{"key":"e_1_2_9_18_1","doi-asserted-by":"publisher","DOI":"10.1021\/acs.jmedchem.8b01723"},{"key":"e_1_2_9_19_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbior.2016.10.002"},{"key":"e_1_2_9_20_1","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2015.00282"},{"key":"e_1_2_9_21_1","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.00377-16"},{"key":"e_1_2_9_22_1","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2015.00282"},{"key":"e_1_2_9_23_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-009-1860-8"},{"key":"e_1_2_9_24_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00253-009-1860-8"},{"key":"e_1_2_9_25_1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkab251"},{"key":"e_1_2_9_26_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi061637j"},{"key":"e_1_2_9_27_1","doi-asserted-by":"publisher","DOI":"10.1038\/378416a0"},{"key":"e_1_2_9_28_1","doi-asserted-by":"publisher","DOI":"10.1021\/acsinfecdis.0c00503"},{"key":"e_1_2_9_29_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2019.06.019"},{"key":"e_1_2_9_30_1","doi-asserted-by":"publisher","DOI":"10.1128\/mBio.02070-15"},{"key":"e_1_2_9_31_1","doi-asserted-by":"publisher","DOI":"10.3389\/fphar.2019.00818"},{"key":"e_1_2_9_32_1","doi-asserted-by":"publisher","DOI":"10.1021\/cr010182v"},{"key":"e_1_2_9_33_1","doi-asserted-by":"publisher","DOI":"10.1128\/mBio.00537-12"},{"key":"e_1_2_9_34_1","doi-asserted-by":"publisher","DOI":"10.1128\/Spectrum.00804-21"},{"key":"e_1_2_9_35_1","doi-asserted-by":"publisher","DOI":"10.1038\/nature25979"},{"key":"e_1_2_9_36_1","volume":"9","author":"Wang S.","year":"2018","journal-title":"Front. 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