RecA polymerization on double-stranded DNA by using single-molecule manipulation: the role of ATP hydrolysis

Abstract

The polymerization of RecA on individual double-stranded DNA molecules is studied. A linear DNA (λ DNA, 48.5 Kb), anchored at one end to a cover glass and at the other end to an optically trapped 3-μm diameter polystyrene bead, serves as a template. The elongation caused by RecA assembly is measured in the presence of ATP and ATP[γS]. By using force extension and hydrodynamic recoil, a value of the persistence length of the RecA-DNA complex is obtained. In the presence of ATP, the polymer length is unstable, first growing to saturation and then decreasing. This suggests a transient dynamics of association and dissociation for RecA on a double-stranded DNA, the process being controlled by ATP hydrolysis. Part of this dynamics is suppressed in the presence of ATP[γS], leading to a stabilized RecA-DNA complex. A one-dimensional nucleation and growth model is presented that may account for the protein assembly.