Figure 4.

Each of the tmaDnaA boxes 1 to 5 within tmaDOR is important for DUE unwinding. Open complex formation by wildtype (WT) tma-oriC and its mutant derivatives (sub1-5), analyzed by P1 assays as described in the legend to Figure 2. A, schematic illustration of the tma-oriC plasmids. Gray bars indicate base substitutions, in which the 12-mer sequence, including the 9-mer tmaDnaA box and its surrounding nucleotides, was replaced by the randomized sequence 5′-CCCAAGCAACAA-3’ (9). The WT sequence is indicated by bold bars. B, representative gel images. C, quantitative data (n = 2) shown as in Figure 2. The mean ± standard deviation activity relative to that of WT at a tmaDnaA concentration of 20 nM are also shown. DOR, DnaA oligomerization region; DUE, duplex unwinding element; tma, Thermotoga maritima.