Fig. 3. The purified factor specifically recognizes the VA1 terminator region. (A) Comparison of the purified factors binding to wild-type and mutant VA1 terminators. The factor purified through the oligo-affinity column was tested for its binding to either wild-type (lanes 1–5 and 11) or mutant (TTTT to ACTG) (lanes 6–10 and 12) VA1 terminator sequence by a gel shift assay in the presence of different concentrations of competitor DNA. The type of competitor and the concentrations(µg/ml) used are marked above the lanes. (B) The purified factor and holo TFIIIC generate identical footprint patterns over the terminator regions (+101 to +232) of the VA1 gene. Immunopurified holo TFIIIC (100 fmol) (lane 3) and 2 (lane 4) or 6 (lane 5) µl of the purified factor from sucrose gradient sedimentation were examined for their footprint on a fragment of the VA1 gene containing both the T1 and T2 terminator regions. The two panels show different exposures of the same gel to show clearly the protection over both terminator regions. The probe was labeled at the 5′ end of the lower strand. Lane 1 is a Maxam–Gilbert G + A reaction and lane 2 is without proteins. The protected sequences and the terminators (T1 and T2) are indicated on the left.