D 10 l of a 50 slurry of Ni2 -NTA-agarose were then added, and the mixture was rotated at 4 for 1 h. Agarose beads were washed with Buffer F (50 mM Tris-HCl (pH 7.5), 300 mM NaCl, 0.2 Tween 20) five times, and DnaK-His and 32 were dissociated from the agarose beads by treating the precipitate with 250 mM imidazole. Proteins were subjected to SDS-PAGE and detected by CBB staining. Experiments u