Schizosaccharomyces pombe Rad22A and Rad22B have similar biochemical properties and form multimeric structures.
The Saccharomyces cerevisiae Rad52 protein has a crucial role in the repair of DNA double-strand breaks by homologous recombination. In vitro, Rad52 displays DNA binding and strand annealing activities and promotes Rad51-mediated strand exchange. Schizosaccharomyces pombe has two Rad52 homologues, Rad22A and Rad22B. Whereas rad22A deficient strains exhibit severe defects in repair and recombination, rad22B mutants have a much less severe phenotype. To better understand the role of Rad22A and Rad22B in double-strand break repair, both proteins were purified to near homogeneity. Using gel retardation and filter binding assays, binding of Rad22A and Rad22B to short single-stranded DNAs was demonstrated. Binding of Rad22A to double-stranded oligonucleotides or linearized plasmid molecules containing blunt ends or short single-stranded overhangs could not be detected. Rad22B also does not bind efficiently to short duplex oligonucleotides but binds readily to DNA fragments containing 3'-overhangs. Rad22A as well as Rad22B efficiently promote annealing of complementary single-stranded DNAs. In the presence of Rad22A annealing of complementary DNAs is almost 90%. Whereas in reactions containing Rad22B the maximum level of annealing is 60%, most likely due to inhibition of the reaction by duplex DNA. Gel-filtration experiments and electron microscopic analyses indicate self-association of Rad22A and Rad22B and the formation of multimeric structures as has been observed for Rad52 in yeast and man.
de Vries
Femke A T
FA
Department of Toxicogenetics, Leiden University Medical Center, PO Box 9600, 2300 RC Leiden, The Netherlands.
Zonneveld
José B M
JB
de Groot
Anton J
AJ
Koning
Roman I
RI
van Zeeland
Albert A
AA
Pastink
Albert
A
D001483:Base Sequence; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008854:Microscopy, Electron; D046912:Multiprotein Complexes; D009154:Mutation; D010641:Phenotype; D012568:Schizosaccharomyces / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D046912:Multiprotein Complexes; C450139:Rad22B protein, S pombe; D029702:Schizosaccharomyces pombe Proteins; C085412:rad52 protein, S pombe