FIG. 4.

(A) The HeLa protein promotes nicking of both flip and flop forms of the hairpin. ³²P-labeled hairpin probes in the flop (lanes 1, 3, and 5) or flip (lanes 2, 4, and 6) orientation were incubated with NS1 alone (lanes 1 and 2) or in the presence of 1 μl (∼0.2 μg) of partially purified HeLa extract (lanes 3 and 4) or with 3 μl (∼0.6 μg) of the same extract (lanes 5 and 6). (B) A hairpin origin with 27 nucleotides of flanking sequence is nicked in the presence of HeLa protein. ³²P-labeled hairpin probes in the flip orientation extending 27 bp beyond the nick site to the DraI site at MVM nucleotide 4896 (P1 in lanes 1 to 3) or 297 bp beyond the nick site to an SspI site at MVM nucleotide 4626 (P in lanes 4 to 6) were incubated with NS1 alone (lanes 2 and 5) or in the presence of 3 μl of partially purified HeLa extract (lanes 3 and 6). C and C1 denote the complexes formed with P and P1, respectively.