Replication protein A physically interacts with the Bloom's syndrome protein and stimulates its helicase activity.
Bloom's syndrome is a rare autosomal recessive disorder characterized by genomic instability and predisposition to cancer. BLM, the gene defective in Bloom's syndrome, encodes a 159-kDa protein possessing DNA-stimulated ATPase and ATP-dependent DNA helicase activities. We have examined mechanistic aspects of the catalytic functions of purified recombinant BLM protein. Through analyzing the effects of different lengths of DNA cofactor on ATPase activity, we provide evidence to suggest that BLM translocates along single-stranded DNA in a processive manner. The helicase reaction catalyzed by BLM protein was examined as a function of duplex DNA length. We show that BLM catalyzes unwinding of short DNA duplexes (</=71 base pairs (bp)) but is severely compromised on longer DNA duplexes (>/=259-bp). The presence of the human single-stranded DNA-binding protein (human replication protein A (hRPA)) stimulates the BLM unwinding reaction on the 259-bp partial duplex DNA substrate. Heterologous single-stranded DNA-binding proteins fail to stimulate similarly the helicase activity of BLM protein. This is the first demonstration of a functional interaction between BLM and another protein. Consistent with a functional interaction between hRPA and the BLM helicase, we demonstrate a direct physical interaction between the two proteins mediated by the 70-kDa subunit of RPA. The interactions between BLM and hRPA suggest that the two proteins function together in vivo to unwind DNA duplexes during replication, recombination, or repair.
Brosh
R M
RM
Laboratory of Molecular Genetics, NIA, National Institutes of Health, Baltimore, Maryland 21224, USA.
Li
J L
JL
Kenny
M K
MK
Karow
J K
JK
Cooper
M P
MP
Kureekattil
R P
RP
Hickson
I D
ID
Bohr
V A
VA
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D001816:Bloom Syndrome; D015153:Blotting, Western; D004247:DNA / Q000378:metabolism; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D006868:Hydrolysis; D009068:Movement; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D053484:RecQ Helicases; D011994:Recombinant Proteins / Q000378:metabolism; D011995:Recombination, Genetic; D051716:Replication Protein A; D012441:Saccharomyces cerevisiae; D013045:Species Specificity
D004268:DNA-Binding Proteins; C497167:RPA1 protein, human; D011994:Recombinant Proteins; D051716:Replication Protein A; D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; C097151:Bloom syndrome protein; D004265:DNA Helicases; D053484:RecQ Helicases