Association of 45- and 34-kDa subunits with VITF-3 activity during purification. Soluble proteins from E. coli coexpressing the A8R and HisA23R proteins were purified by column chromatography. At each chromatographic step, fractions were tested for intermediate transcription factor activity and active fractions were pooled. (A) Total soluble protein (Sol) and pooled active fractions from Talon, HQ, and heparin (Hep) columns were analyzed by SDS/PAGE and silver staining. The positions of protein markers (M) are shown on the right. (B) Pooled fractions in A were tested for intermediate transcription factor activity as described in the legend to Fig. 1. Additional lanes: B, buffer control; X, VITF-X.