Figure 2.

Reactivity of γ ori DNA to KMnO₄ in the presence of His-tagged, copy-up π protein and increasing concentrations of ATP or AMP-PCP. pMF36 DNA template was incubated with 1.00 µg of the copy-up variant protein and with 0, 2, 4, 8, 16 or 32 mM ATP (A), or AMP-PCP (C). For the control reaction, the reactivity of the DNA probe to KMnO₄ in the absence of protein and/or ATP/AMP-PCP was examined. Reactive bases were identified using the DNA sequencing ladder displayed in the last four lanes. The positions of the A + T-rich region [black bar and Chen et al. (9)] and iterons (large, vertical, gray arrow) are indicated. Reaction products were processed as described (13). Autoradiographs were quantified (B and D) as described (13). Quantifications of lanes containing no protein (without ATP or AMP-PCP), protein only (without ATP or AMP-PCP) and protein with ATP or AMP-PCP (32 nM) are presented. Bands of interest (singular or in clusters) are indicated as follows: small black arrows highlight bands of increasing intensity; small gray/white arrows highlight bands of decreasing intensity; black bars indicate decreased band intensities at low nucleotide levels in the presence of Rep protein.