Cooperative binding of Agrobacterium tumefaciens VirE2 protein to single-stranded DNA.
The VirE2 protein of Agrobacterium tumefaciens Ti plasmid pTiA6 is a single-stranded-DNA-binding protein. Density gradient centrifugation studies showed that it exists as a tetramer in solution. Monomeric VirE2 active in DNA binding could also be obtained by using a different protein isolation procedure. VirE2 was found to be thermolabile; brief incubation at 37 degrees C abolished its DNA-binding activity. It was insensitive to the sulfhydryl-specific reagent N-ethylmaleimide. Removal of the carboxy-terminal 37 residues of the 533-residue VirE2 polypeptide led to complete loss of DNA-binding activity; however, chimeric fusion proteins containing up to 125 residues of the VirE2 C terminus were inactive in DNA binding. In nuclease protection studies, VirE2 protected single-stranded DNA against degradation by DNase I. Analysis of the DNA-VirE2 complex by electron microscopy demonstrated that VirE2 coats a single-stranded DNA molecule and that the binding of VirE2 to its substrate is cooperative.
Sen
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Department of Biochemistry, University of Minnesota, St. Paul 55108.
Pazour
G J
GJ
Anderson
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Das
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A
D001426:Bacterial Proteins / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003852:Deoxyribonucleoproteins / Q000648:ultrastructure; D007473:Ion Channels; D008854:Microscopy, Electron; D008970:Molecular Weight; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015183:Restriction Mapping; D012231:Rhizobium / Q000378:metabolism*; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D003852:Deoxyribonucleoproteins; D007473:Ion Channels; D011993:Recombinant Fusion Proteins; C051583:virE2 protein, Agrobacterium