Figure 7.

Constitutive expression of CopM restores both plastocyanin and cellular copper levels. (A) Western blot analysis of plastocyanin levels after copper addition in WT, WTM, COPR, COPRM, and COPB strains. Cells were grown in BG11C-Cu medium to mid-log growth phase and exposed for 24 h to copper 1 μmol/L. Five micrograms of total protein from soluble extracts was separated by 15% SDS-PAGE and analyzed by western blot to detect plastocyanin. (B) Quantification of plastocyanin levels in response to copper addition in WT (white circles), WTM (black circles), COPR (white squares), COPRM (black squares), and COPB (white triangles) strains. Western blot signal of three independent experiments was quantified using ImageJ program. Plastocyanin levels were normalized to the GSI signal. Error bars represent SE. (C) Total intracellular copper contents in WT, WTM, COPR, COPRM, and COPB strains. Cells were grown in BG11C-Cu medium to mid-log growth phase and exposed for 1 μmol/L of copper for 5 h. Cells were centrifuged, washed twice with BG11C-Cu, and dried. Hundred micrograms of dried cells was microwave digested, dissolved in suprapure HNO₃, and analyzed by ICP. Error bars represent SE from three independent experiments.