Mass spectrometry analysis of Flag-H9N2-NS2 interacting candidates

To identify SUMOylated factors that interact with NS2 via SIM-SUMO interaction, FLAG-tagged H9N2-NS2 was co-transfected into HEK293T cells together with plasmids encoding His-SUMO1 and Myc-Ubc9. After transfection for 24 hours, cells were infected with H9N2 AIV for a further 24 hours and Flag-NS2 was purified using anti-Flag beads. Purified protein complexes were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). To identify SUMOylated factors that interact with NS2 via SIM-SUMO interaction, FLAG-tagged H9N2-NS2 was co-transfected into HEK293T cells together with plasmids encoding His-SUMO1 and Myc-Ubc9. After transfection for 24 hours, cells were infected with H9N2 AIV for a further 24 hours and Flag-NS2 was purified using anti-Flag beads. Purified protein complexes were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS).