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Successive mutation of the glutamic acids in p6 gradually increases HIV-1 Gag ubiquitination. (A,B) HeLa cells were co-transfected with HA-tagged ubiquitin and pNLenv1 expression plasmids as indicated. Gag was immunoprecipitated from whole cell lysates using anti-HIV antibodies and Gag ubiquitination was detected by western blot stained for HA. The amount of precipitated Gag was detected by anti-CA staining.
