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![[Figure 6]](cb5130fig6mag.jpg)
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Figure 6
Cleavage of a DNA molecule in the form of a Holliday junction by the Hjc_15-6 enzyme. Lane 1 contains intact X-shaped dsDNA. The resolving enzyme activity of Hjc_15-6 is visualized in lanes 2–5 after incubation for 30 min at 50°C. The reaction mixtures in lanes 2–5 contained 10 pmol X-shaped substrate DNA, 10 mM Tris–HCl pH 8.5, 100 mM KCl, 10 mM MgCl2 and 35 pmol purified Hjc_15-6 expressed in complex (LB) medium. Lanes 3 and 5 contained an additional 5 mM ATP, whereas the samples in lanes 2 and 4 were not treated with ATP. In lanes 4 and 5, the Hjc_15-6 used had been purified by dual chromatography to remove contaminants. In lane 6, 0.1 µg double-stranded blunt-end λ/HindIII DNA fragments were used as a control. In lane 7, a reaction mixture with 1 µg blunt-end λ/HindIII and 1.3 µg (approximately 35 pmol) Hjc_15-6 (purified twice) was applied after incubation for 30 min at 37°C. Except for the substrate DNA (1 µg blunt-end λ/HindIII) the reaction mixture is the same as in lane 4, indicating that Hjc_15-6 has no nuclease activity towards the nonbranched blunt-end dsDNA. S and P indicate the positions of substrate and product, respectively. |
![[Figure 6]](cb5130fig6.jpg)
 | STRUCTURAL BIOLOGY |
ISSN: 2059-7983
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