Magnesium Ion-dependent Activation of the RecA Protein Involves the C Terminus

Optimal conditions for RecA protein-mediated DNA strand exchange include 6–8 mm Mg2+ in excess of that required to form complexes with ATP. We provide evidence that the free magnesium ion is required to mediate a conformational change in the RecA protein C terminus that activates RecA-mediated DNA strand exchange. In particular, a “closed” (low Mg2+) conformation of a RecA nucleoprotein filament restricts DNA pairing by incoming duplex DNA, although single-stranded overhangs at the ends of a duplex allow limited DNA pairing to occur. The addition of excess Mg2+ results in an “open” conformation, which can promote efficient DNA pairing and strand exchange regardless of DNA end structure. The removal of 17 amino acid residues at theEscherichia coli RecA C terminus eliminates a measurable requirement for excess Mg2+ and permits efficient DNA pairing and exchange similar to that seen with the wild-type protein at high Mg2+ levels. Thus, the RecA C terminus imposes the need for the high magnesium ion concentrations requisite in RecA reactions in vitro. We propose that the C terminus acts as a regulatory switch, modulating the access of double-stranded DNA to the presynaptic filament and thereby inhibiting homologous DNA pairing and strand exchange at low magnesium ion concentrations.