Figure 3.

Hel308 from archaea preferentially targets fork DNA for unwinding. (A) Gel-retardation binding assays of Hel308a on the substrates in Figure 2E in 1 mM magnesium. Reactions were at 45°C for 10 min and contained 2 nM DNA substrate mixed with 0, 1, 2, 10, 50 and 100 nM Hel308a. (B) The same reactions as in (A), but containing an additional 1 mM ATP in the gel, and all buffers. (C). Unwinding reactions of Hel308a on flayed duplex (lanes a–f), fork with leading strand only (lanes g–l), fork with lagging strand only (fork-2, lanes m–r), fork with both leading and lagging strands (fork-1, lanes s–x) and Holliday junction (lanes y–dd). Reactions were for 20 min at 45°C containing 2 nM DNA, 5 mM MgCl₂, 5 mM ATP and zero (lanes a, g, m, s and y) or 1, 5, 10, 25 and 50 nM Hel308a.