Figure 7.

Effect of ATP, AMPPCP, and AMPPNP on the digestion pattern of T. maritima CopA WT, ΔNMBD, and CXXC and H479Q mutant proteins. SDS−PAGE was performed as described in Experimental Procedures. Purified CopA protein (0.5 mg/mL) was incubated with 0.5 mg/mL papain at 50 °C in the absence and presence of nucleotides. (a and c) Time course digestion experiments with CopA WT (a) and H479Q (c) were performed in the absence of copper (1 mM BCS present). Nucleotides were added as indicated in the figure. The time of digestion was 10 min for lanes 2, 5, 8, and 11; 20 min for lanes 3, 6, 9, and 12; and 30 min for lanes 4, 7, 10, and 13. (b) Digestion of CopA WT, ΔNMBD, and CXXC was performed for 30 min in the presence of either 10 μM CuCl₂ (+) or 1 mM BCS (−), with or without 2 mM ATP or 2 mM AMPPCP as indicated.