The C-terminus of the E. coli RecA protein modulates the DNA binding competition with SSB.
Eggler AL, Lusetti SL, Cox MM
Journal of Biological Chemistry (2003)
Category: DNA repair ¤ Added: Feb 25, 2003 ¤ Rating: ◊◊
The nucleation step of Escherichia coli RecA filament formation on single-stranded DNA (ssDNA) is strongly inhibited by prebound E. coli ssDNA binding protein (SSB). The capacity of RecA protein to displace SSB is dramatically enhanced in RecA proteins with C-terminal deletions. The displacement of SSB by RecA protein is progressively improved when 6, 13, and 17 C-terminal amino acids are removed from the RecA protein relative to the full-length protein. The C-terminal deletion mutants also more readily displace yeast Replication Protein A (RPA) than does the full-length protein. Thus, the RecA protein has an inherent and robust capacity to displace SSB from ssDNA. However, the displacement function is suppressed by the RecA C-terminus, providing another example of a RecA activity with C-terminal modulation. RecA*C17 also has an enhanced capacity relative to wild-type RecA protein to bind ssDNA containing secondary structure. Added Mg(2+) enhances the ability of wild-type RecA and the RecA C-terminal deletion mutants to compete with SSB and RPA. The overall binding of RecA*C17 mutant protein to linear ssDNA is increased further by the mutation E38K, previously shown to enhance SSB displacement from ssDNA. The double mutant RecA*C17/E38K displaces SSB somewhat better than either individual mutant protein under some conditions, and exhibits a higher steady-state level of binding to linear ssDNA under all conditions.