FIGURE 1.
Yku binds to duplex and tail-duplex DNA substrates formed by telomeric DNA. A, purification of Ku from yeast. A TAP-tagged Yku80 was employed to purify Yku from yeast BJ2168. A Coomassie Blue-stained 10% SDS-polyacrylamide gel is given. Lane 1 (M) shows the molecular weight marker; lane 2 shows 1 μg of purified Yku. B, Yku binds to tailed-duplex DNA formed by telomeric DNA. ∼2 ng of 32P-radiolabeled DNA formed by T1T2 (left panel) or T3T4 (right panel) was incubated with 50 nm (lanes 2), 150 nm (lanes 3), or 450 nm (lanes 4) of purified Yku at room temperature for 10 min and then analyzed by a 6% polyacrylamide gel. An autoradiogram is shown here. C, DNase I footprints of Yku to tailed-duplex DNA. The footprint was done using a tailed-duplex DNA substrate formed by RT1 and RT2. Five nm each of the DNA substrates were used in the reactions. The Yku concentration was 7, 66, 133, and 200 nm, respectively. The autoradiograms of DNase I footprints (left) and the DNA sequences of tailed-duplex DNA (right) are shown. The regions protected from DNase I digestion are bracketed.