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Nakamura, T. ; Ito, J. ; Mourao, A. ; Wahida, A. ; Nakagawa, K.* ; Mishima, E. ; Conrad, M.

A tangible method to assess native ferroptosis suppressor activity.

Cell Rep. Methods 4, 18:100710 (2024)
Verlagsversion DOI PMC
Open Access Gold
Creative Commons Lizenzvertrag
Ferroptosis, a regulated cell death hallmarked by unrestrained lipid peroxidation, plays a pivotal role in the pathophysiology of various diseases, making it a promising therapeutic target. Glutathione peroxidase 4 (GPX4) prevents ferroptosis by reducing (phospho)lipid hydroperoxides, yet evaluation of its actual activity has remained arduous. Here, we present a tangible method using affinity-purified GPX4 to capture a snapshot of its native activity. Next to measuring GPX4 activity, this improved method allows for the investigation of mutational GPX4 activity, exemplified by the GPX4U46C mutant lacking selenocysteine at its active site, as well as the evaluation of GPX4 inhibitors, such as RSL3, as a showcase. Furthermore, we apply this method to the second ferroptosis guardian, ferroptosis suppressor protein 1, to validate the newly identified ferroptosis inhibitor WIN62577. Together, these methods open up opportunities for evaluating alternative ferroptosis suppression mechanisms.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Korrespondenzautor
Schlagwörter Cp: Molecular Biology ; Fsp1 ; Gpx4 ; Lc-ms/ms ; Rsl3 ; Affinity Purification ; Biochemistry ; Cell Death ; Drug Discovery ; Enzyme Assay ; Lipid Peroxidation ; Pull-down Assay ; Selenocysteine; Cancer-cells; Separation; Death; Gpx4
ISSN (print) / ISBN 2667-2375
e-ISSN 2667-2375
Zeitschrift Cell Reports Methods
Quellenangaben Band: 4, Heft: 3, Seiten: 18, Artikelnummer: 100710 Supplement: ,
Verlag Elsevier
Verlagsort 50 Hampshire St, Floor 5, Cambridge, Ma 02139 Usa
Nichtpatentliteratur Publikationen
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Metabolism and Cell Death (MCD)
Institute of Structural Biology (STB)
Förderungen JSPS KAKENHI
CRC TRR 353
German Federal Ministry of Education and Research (BMBF) FERROPATH
European Research Council (ERC)
Deutsche Forschungsgemeinschaft (DFG)