Figure 4.

Overexpression and calcium binding of recombinant calsymin in E. coli. Coomassie brilliant blue staining (A) and autoradiograph (B) after incubation with ⁴⁵Ca²⁺, of protein profiles immobilized on a PVDF membrane. Crude protein extracts were obtained from the culture supernatant (lanes 1 and 2) or from the cell pellet (lanes 3 and 4). Exponential cell cultures were either uninduced (lanes 2 and 4) or induced with 0.2% arabinose for 6 h (lanes 1 and 3) before harvesting. The recombinant protein was extracted under denaturing conditions from a crude protein extract of E. coli Top10 cells (Invitrogen) induced with arabinose and purified by Ni-chelation chromatography (lane 5). The calsymin band is marked with an arrowhead.