A novel site-specific recombinase encoded by the Streptococcus pyogenes plasmid pSM19035.
Genetic evidence suggests that the beta protein encoded by the Streptococcus pyogenes plasmid pSM19035 is involved both in the resolution of plasmid multimers into monomers and in DNA inversion. In this report we show that the highly purified beta protein is unable to mediate DNA recombination unless a host factor(s) is provided. In the presence of the host factor(s), the beta protein is able to catalyze in vitra intramolecular recombination between two specific sites on supercoiled templates: DNA resolution was obtained when the two recombination sites were directly oriented, whereas DNA inversion was the product if the recombination sites were in inverse orientation. In the absence of the host factor(s) the beta protein forms a specific complex with its target site. The beta protein binding site has been localized by DNase I footprinting to an 85 bp region that can be divided into two discrete sites (I and II). These sites are about 34 bp in length, they are separated by about 16 bp, and contain two 12 to 13 bp imperfectly conserved sequences (half-sites) with dyad axis symmetry. The protein binds co-operatively to sites I and II; between 3.6 and 4.2 beta protein protomers are required to saturate the DNA substrate. These data, together with gel retardation assays, suggest that the protein binds to DNA as two dimers, one to each discrete site, and that the dimers probably interact with each other. The beta protein binding site, though it resembles that of other DNA resolvases of the Tn3/gamma delta (Tn1000) family, differs in that only two adjacent sites are found (sites I and II), while those DNA resolvases normally bind to three adjacent sites. The results presented here suggest that a host factor(s) could work as an accessory effector to compensate for the absence of the third binding site.
Rojo
F
F
Centro de Biología Molecular Severo Ochoa, C.S.I.C., Universidad Autónoma de Madrid, Spain.
Alonso
J C
JC
D001426:Bacterial Proteins / Q000378:metabolism; D001483:Base Sequence; D004254:DNA Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000378:metabolism; D010957:Plasmids / Q000235:genetics*; D013297:Streptococcus pyogenes / Q000235:genetics*; D013379:Substrate Specificity; D019895:Transposases
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D004254:DNA Nucleotidyltransferases; C056830:DNA invertase Gin; D009713:Nucleotidyltransferases; D019895:Transposases