Bacillus subtilis Bacteriophage SPP1 G40P Helicase Lacking the N-terminal Domain Unwinds DNA Bidirectionally

Bacillus subtilis bacteriophage SPP1 G40P hexameric replicative DNA helicase unidirectionally translocates with a 5′→3′ polarity while separating the DNA strands. A G40P mutant derivative lacking the N-terminal domain (containing amino acid residues 110–442 from G40P, G40PΔN109) was purified and characterized. G40PΔN109 showed an ATPase activity that was dependent on the presence of single-stranded (ss) DNA. Unlike G40P, G40PΔN109 was shown to bind with similar affinity both ssDNA arms of forked structures by nuclease protection assays. In a pH-dependent manner, G40PΔN109 unwound a branched double-arm substrate preferentially with a 3′→5′ polarity. Our results show that the linker region and the C-terminal domain of G40P are sufficient to render an enzyme capable of encircling the ssDNA tails of the forked DNA and to unwind DNA with both 5′→3′ and 3′→5′ polarity. The presence of the N-terminal domain, which does not play an essential role in helicase action, might be required indirectly for strand discrimination and polarity of translocation.