Interaction of the Gifsy-1 Xis protein with the Gifsy-1 attP sequence.
The Gifsy-1 phage integrates site specifically into the Salmonella chromosome via an integrase-mediated site-specific recombination mechanism. Initial genetic analysis suggests that Gifsy-1 integrase-mediated excision of the Gifsy-1 phage is influenced by proteins encoded by both the Gifsy-1 and the Gifsy-2 phages. Our studies show that the Gifsy-1 Xis protein regulates the directionality of integrase-mediated excision of the Gifsy-1 phage. Electrophoretic mobility shift assays, DNase I footprinting, dimethyl sulfate (DMS) interference assays, and DMS protection assays were used to identify a 31-base-pair sequence in the attP region to which the Gifsy-1 protein binds. The results suggest that this recombination directionality factor binds in vitro to three imperfect direct repeats, spaced 10 base pairs apart, in a sequential and cooperative manner in the absence of other phage-encoded proteins. Our studies suggest that, while the Gifsy-1 Xis does not require additional factors for specific and high-affinity binding, it may form a microfilament on DNA similar to that described for the phage lambda Xis protein.
Flanigan
Asa
A
Department of Microbiology, University of Illinois at Urbana-Champaign, 601 South Goodwin Avenue, Urbana, IL 61801, USA.
Gardner
Jeffrey F
JF
D000595:Amino Acid Sequence; D001287:Attachment Sites, Microbiological; D001483:Base Sequence; D018983:DNA Footprinting; D004269:DNA, Bacterial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D008969:Molecular Sequence Data; D011485:Protein Binding; D012483:Salmonella Phages / Q000235:genetics* / Q000378:metabolism; D012486:Salmonella typhimurium / Q000235:genetics / Q000821:virology*; D013463:Sulfuric Acid Esters / Q000378:metabolism; D014764:Viral Proteins / Q000378:metabolism*
D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D013463:Sulfuric Acid Esters; D014764:Viral Proteins; C007482:dimethyl sulfate