title: Purification and characterization of legionella U-box-type E3 ubiquitin ligase.
abstract: Bacterial virulence proteins often mimic host eukaryotic proteins to modify or disturb host cellular -pathways. Increasing lines of evidence show that many bacterial effector proteins have E3 ubiquitin ligase activity. The effector protein LubX is one such bacterial E3 ubiquitin ligase. We describe here the method to purify soluble LubX protein using GST-tag and Escherichia coli overexpression systems. Using the purified protein together with recombinant ubiquitin, E1, and E2 enzymes, ubiquitin ligase activity is analyzed by the in vitro ubiquitination assay.
authors.affiliation: Research Institute for Microbial Diseases, Osaka University, Osaka, Japan. hnagai@biken.osaka-u.ac.jp
authors.forename: Hiroki
authors.initials: H
authors.lastname: Nagai
authors.forename: Tomoko
authors.initials: T
authors.lastname: Kubori
mesh_terms: D001426:Bacterial Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D004789:Enzyme Activation; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D007875:Legionella / Q000201:enzymology* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000302:isolation & purification* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D044767:Ubiquitin-Protein Ligases