FIGURE 4.

N-term MBD mutants exhibit basal phosphorylation and not copper-dependent hyperphosphorylation. A, schematic showing N-terminal MBD mutants (right). YST cells were co-transfected with pTyrosinase alone (Tyr) or with various ATP7B constructs as indicated. The black reaction product indicates the presence of copper-dependent tyrosinase activity (left). B and C, GFP-WTATP7B and the two N-terminal mutants were expressed and evaluated for phosphorylation as described under “Experimental Procedures.” B, autoradiogram and blot showing ³²P incorporation and protein levels of the three constructs. C, bar graph showing the mean relative specific phosphorylation of GFP-WTATP7B, 1–63 Δ1–4MBD, and 1–63 Δ1–4MBD C>S normalized to the −copper (−Cu) condition. n = 4. Error bars, mean ± S.E.; *, p < 0.05 using two-tailed t test; NS, not significant.