Purification of VP19C and the DNA-binding property of the purified protein.

<p>A. Expression and purification of VP19C protein from yeast cells. Yeast cells induced for expression of GST-VP19C and the proteins purified from these cells were analyzed by SDS/PAGE using 4-12% Nu-PAGE gels and detected by Coomassie Blue staining. Total proteins extracted from uninduced cultures (Un) and cells induced (In) with glucose are shown. The protein fractions, flow-through (FT) and wash (Wash) obtained during the purification are also shown. GST-VP19C bound with glutathione–Sepharose beads before elution (Beads) and the eluted purified protein (Elution) were detected. Molecular-mass markers; lane 2. B. VP19C specifies a DNA-binding property. In the EMSA a small dsDNA (66 bp) radiolabeled probe (1.5 ng) was incubated with 1.5 µg (lane 2) and 3 µg VP19C (lanes 3 and 4) or no protein (lane 1). In lane 4, a 50-fold excess of non-labeled DNA was added to the VP19C-DNA composition. The nucleo-protein complex (NPC) and the location of the free probe are indicated on the left of the panel.