Skip to main content
. 2013 Jun 21;288(31):22809–22820. doi: 10.1074/jbc.M113.482091

FIGURE 4.

FIGURE 4.

Point-mutated DJ-1 metal binding analysis. A, Western blot (WB) analysis, using anti-His6- and anti-DJ-1-specific antibodies of purified recombinant human DJ-1 WT and point-mutated DJ-1 A104T, DJ-1 D149A, DJ-1 C106A, and DJ-1 WT-His6 tag as positive control. B, AAS analysis of total amount of copper bound to recombinant human DJ-1 proteins, as shown in A, after 24 h exposure time. Statistically significant changes in total copper binding compared with WT DJ-1 are indicated with asterisks, **, p < 0.01; ***, p < 0.001. C, initial binding affinities for metal binding to DJ-1 recombinant human DJ-1 proteins, as shown in A. Calculated average MST dissociation constants (KD) for recombinant human DJ-1 WT and point-mutated variants using Cu(I), Cu(II), and Hg(II) as ligands. Lack of detected binding indicated with − sign. D, crystal structure model of dimeric human DJ-1 with the studied mutation sites indicated with *, and potential metal binding areas indicated within squares (dotted line). Protein structure model was adopted from Protein Data Bank code 2R1V.