Complete inhibition of Streptococcus pneumoniae RecA protein-catalyzed ATP hydrolysis by single-stranded DNA-binding protein (SSB protein): implications for the mechanism of SSB protein-stimulated DNA strand exchange.
The ATP-dependent three-strand exchange activity of the Streptococcus pneumoniae RecA protein (RecA(Sp)), like that of the Escherichia coli RecA protein (RecA(Ec)), is strongly stimulated by the single-stranded DNA-binding protein (SSB) from either E. coli (SSB(Ec)) or S. pneumoniae (SSB(Sp)). The RecA(Sp) protein differs from the RecA(Ec) protein, however, in that its ssDNA-dependent ATP hydrolysis activity is completely inhibited by SSB(Ec) or SSB(Sp) protein, apparently because these proteins displace RecA(Sp) protein from ssDNA. These results indicate that in contrast to the mechanism that has been established for the RecA(Ec) protein, SSB protein does not stimulate the RecA(Sp) protein-promoted strand exchange reaction by facilitating the formation of a presynaptic complex between the RecA(Sp) protein and the ssDNA substrate. In addition to acting presynaptically, however, it has been proposed that SSB(Ec) protein also stimulates the RecA(Ec) protein strand exchange reaction postsynaptically, by binding to the displaced single strand that is generated when the ssDNA substrate invades the homologous linear dsDNA. In the RecA(Sp) protein-promoted reaction, the stimulatory effect of SSB protein may be due entirely to this postsynaptic mechanism. The competing displacement of RecA(Sp) protein from the ssDNA substrate by SSB protein, however, appears to limit the efficiency of the strand exchange reaction (especially at high SSB protein concentrations or when SSB protein is added to the ssDNA before RecA(Sp) protein) relative to that observed under the same conditions with the RecA(Ec) protein.
Steffen
Scott E
SE
Department of Biochemistry, The Johns Hopkins University, Bloomberg School of Public Health, Baltimore, Maryland 21205, USA.
Katz
Francine S
FS
Bryant
Floyd R
FR
D000255:Adenosine Triphosphate / Q000378:metabolism*; D002384:Catalysis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D011938:Rec A Recombinases / Q000037:antagonists & inhibitors*; D013296:Streptococcus pneumoniae / Q000378:metabolism*
D004268:DNA-Binding Proteins; D000255:Adenosine Triphosphate; D011938:Rec A Recombinases