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. 2011 Feb 25;286(16):14019–14027. doi: 10.1074/jbc.M110.190710

FIGURE 2.

FIGURE 2.

OA-NO2 reacts with Keap1 in a cellular milieu. HEK-293T cells were transfected with FLAG-CMV (empty vector) or FLAG-Keap1-overexpressing vector and treated with the indicated concentrations of OA-NO2. A, Keap1-adducted OA-NO2 was exchanged to β-ME from immunoprecipitated Keap1 in the presence of a [13C18]OA-NO2 internal standard and quantified by LC-MS/MS as β-ME-OA-NO2. The lower panels show transfection efficiency of FLAG-Keap1 constructs. B, β-ME-OA-NO2 levels captured upon exchange to β-ME in immunoprecipitated WT Keap1 and the following Cys to Ser mutant Keap1: C273S, C288S, C273S/C288S, or C151S/C273S/C288S. β-ME exchange reactions were conducted in the presence of [13C18]OA-NO2 and quantified by LC-MS/MS. Values are represented as mean ± S.D. (error bars) *, p < 0.05; **, p < 0.01; ***, p < 0.001 when compared with respective control. ns, not significant.