Real-time observation of bacteriophage T4 gp41 helicase reveals an unwinding mechanism

Publication Type:

Journal Article

Source:

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, NATL ACAD SCIENCES, Volume 104, Number 50, 2101 CONSTITUTION AVE NW, WASHINGTON, DC 20418 USA, p.19790-19795 (2007)

DOI:

10.1073/pnas.0709793104

Keywords:

FORCE; GENE-41 PROTEIN; HEXAMERIC HELICASE; MOLECULE; NS3 HELICASE; PROCESSIVITY; REPLICATION; SINGLE-STRANDED-DNA; T4; TRANSLOCATION

Abstract:

Helicases are enzymes that couple ATP hydrolysis to the unwinding of double-stranded (ds) nucleic acids. The bacteriophage T4 helicase (gp41) is a hexameric helicase that promotes DNA replication within a highly coordinated protein complex termed the replisome. Despite recent progress, the gp41 unwinding mechanism and regulatory interactions within the replisome remain unclear. Here we use a single tethered DNA hairpin as a real-time reporter of gp41-mediated dsDNA unwinding and single-stranded (ss) DNA translocation with 3-base pair (bp) resolution. Although gp41 translocates on ssDNA as fast as the in vivo replication fork (approximate to 400 bp/s), its unwinding rate extrapolated to zero force is much slower (approximate to 30 bp/s). Together, our results have two implications: first, gp41 unwinds DNA through a passive mechanism; second, this weak helicase cannot efficiently unwind the T4 genome alone. Our results suggest that important regulations occur within the replisome to achieve rapid and processive replication.