Visualisation of human rad52 protein and its complexes with hRad51 and DNA.
The human Rad52 protein stimulates joint molecule formation by hRad51, a homologue of Escherichia coli RecA protein. Electron microscopic analysis of hRad52 shows that it self-associates to form ring structures with a diameter of approximately 10 nm. Each ring contains a hole at its centre. hRad52 binds to single and double-stranded DNA. In the ssDNA-hRad52 complexes, hRad52 was distributed along the length of the DNA, which exhibited a characteristic "beads on a string" appearance. At higher concentrations of hRad52, "super-rings" (approximately 30 nm) were observed and the ssDNA was collapsed upon itself. In contrast, in dsDNA-hRad52 complexes, some regions of the DNA remained protein-free while others, containing hRad52, interacted to form large protein-DNA networks. Saturating concentrations of hRad51 displaced hRad52 from ssDNA, whereas dsDNA-Rad52 complexes (networks) were more resistant to hRad51 invasion and nucleoprotein filament formation. When Rad52-Rad51-DNA complexes were probed with gold-conjugated hRad52 antibodies, the presence of globular hRad52 structures within the Rad51 nucleoprotein filament was observed. These data provide the first direct visualisation of protein-DNA complexes formed by the human Rad51 and Rad52 recombination/repair proteins.
Van Dyck
E
E
Clare Hall Laboratories, Imperial Cancer Research Fund, South Mimms, Hertfordshire, EN6 3LD, UK.
Hajibagheri
N M
NM
Stasiak
A
A
West
S C
SC
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D004247:DNA / Q000737:chemistry* / Q000648:ultrastructure; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000648:ultrastructure; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure; D006801:Humans; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D011485:Protein Binding; D011487:Protein Conformation; D051135:Rad51 Recombinase; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D018411:Spodoptera
D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D004247:DNA; C495153:RAD51 protein, human; D051135:Rad51 Recombinase