MBP- Pf Ctr369Nt-Sbinds copper in vitro and within E. coli host cells. A. Affinity-purified MBP-Pf Ctr369Nt-S or MBP was incubated with CuCl2 in the presence (+) or absence (−) of ascorbic acid in vitro. The BCA release assay detected copper with (solid bars) or without (open bars) the addition of ascorbic acid. The concentration of the copper standard (CuCl2 only) was equimolar to the amount of protein used. B. CuCl2 was added to the E. coli cell growth medium after the induction of recombinant protein expression. Following affinity purification, protein-bound copper was detected using the BCA release assay with (solid bars) or without (open bars) the addition of ascorbic acid. The addition of copper to the E. coli growth medium increased recombinant protein yield (inset). In all panels, MBP serves as a control and * represents statistical significance (p < 0.05). Results are means ± S.D. of triplicate measurements done for duplicate dialysis sacs.