{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,5]],"date-time":"2026-06-05T06:14:35Z","timestamp":1780640075065,"version":"3.54.1"},"reference-count":68,"publisher":"Wiley","issue":"21","license":[{"start":{"date-parts":[[2025,9,29]],"date-time":"2025-09-29T00:00:00Z","timestamp":1759104000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000781","name":"ERC","doi-asserted-by":"publisher","award":["101087582"],"award-info":[{"award-number":["101087582"]}],"id":[{"id":"10.13039\/501100000781","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["chemistry-europe.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["ChemBioChem"],"published-print":{"date-parts":[[2025,11,8]]},"abstract":"<jats:p>\n                    Ubiquitination of proteins is one of the most crucial post\u2010translational modifications in eukaryotic cells, typically involving conjugation of ubiquitin to a lysine residue in a substrate using a three\u2010enzyme cascade that relies on ATP as energy source. The pathogen\n                    <jats:italic>Legionella pneumophila<\/jats:italic>\n                    , in contrast, employs a totally divergent pathway to ubiquitinate cellular host proteins in an unconventional manner that is crucial for bacterial proliferation. This multistep process is orchestrated by effector proteins from the SidE family which initially use NAD\n                    <jats:sup>+<\/jats:sup>\n                    to adenosine diphosphate (ADP)\u2010ribosylate ubiquitin in a mono\u2010ADP\u2010ribosyltransferase (mART) domain. The subsequent step relies on SidE phosphodiesterase activity to conjugate phosphoribosyl\u2010Ubiquitin to serine residues in host substrates. Through these phosphoribosyl\u2010ubiquitinating events,\n                    <jats:italic>Legionella<\/jats:italic>\n                    is able to gain local control over the host's ubiquitin system and simultaneously evades immune responses. Hence, pursuing new inhibitors which can disrupt these crucial steps in bacterial infection are essential towards further understanding and potentially blocking\n                    <jats:italic>Legionella<\/jats:italic>\n                    infection. Here, we present the application of an\n                    <jats:italic>\u03b5<\/jats:italic>\n                    \u2010NAD\n                    <jats:sup>+<\/jats:sup>\n                    consumption\u2010based fluorogenic assay to identify small molecule modulators of the SdeA effector enzyme in a High\u2010Throughput Screening format, where over 600 compounds were screened. As a result, a potent inhibitor named cephalosporin C Zn\n                    <jats:sup>2+<\/jats:sup>\n                    salt was discovered showing an IC\n                    <jats:sub>50<\/jats:sub>\n                    of 221\u2009nM. To investigate the inhibitory properties more deeply, various cephalosporin analogs were synthesized where variations in charge and carbon length were introduced and their inhibitory efficiencies measured and compared. Our findings suggest that the inhibition is primarily attributed to the presence of the Zn\n                    <jats:sup>2+<\/jats:sup>\n                    ion, rather than the cephalosporin core. We next compared the inhibitory potential of other bivalent metal ions, illustrating that the zinc ion causes the best inhibition of the\n                    <jats:italic>Legionella<\/jats:italic>\n                    effector.\n                  <\/jats:p>","DOI":"10.1002\/cbic.202500513","type":"journal-article","created":{"date-parts":[[2025,9,29]],"date-time":"2025-09-29T10:19:42Z","timestamp":1759141182000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Development and Validation of a High\u2010Throughput Screening Assay for the Legionella ADP\u2010Ribosyl Transferase SdeA"],"prefix":"10.1002","volume":"26","author":[{"given":"Halana C.","family":"Vlaming","sequence":"first","affiliation":[{"name":"Leiden University Medical Centre Dept. 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