Functional and physical interaction between WRN helicase and human replication protein A.
The human premature aging disorder Werner syndrome (WS) is associated with a large number of symptoms displayed in normal aging. The WRN gene product, a DNA helicase, has been previously shown to unwind short DNA duplexes (</=53 base pairs) in a reaction stimulated by single-stranded DNA-binding proteins. We have studied the helicase activity of purified WRN protein on a variety of DNA duplex substrates to characterize the unwinding properties of the enzyme in greater detail. WRN helicase can catalyze unwinding of long duplex DNA substrates up to 849 base pairs in a reaction dependent on human replication protein A (hRPA). Escherichia coli SSB and bacteriophage T4 gene 32 protein (gp32) completely failed to stimulate WRN helicase to unwind long DNA duplexes indicating a specific functional interaction between WRN and hRPA. So far, there have been no reports of any physical interactions between WRN helicase and other proteins. In support of the functional interaction, we demonstrate a direct interaction between WRN and hRPA by coimmunoprecipitation of purified proteins. The physical and functional interaction between WRN and hRPA suggests that the two proteins may function together in vivo in a pathway of DNA metabolism such as replication, recombination, or repair.
Brosh
R M
RM
Laboratory of Molecular Genetics, NIA, National Institutes of Health, Baltimore, Maryland 21224, USA.
Orren
D K
DK
Nehlin
J O
JO
Ravn
P H
PH
Kenny
M K
MK
Machwe
A
A
Bohr
V A
VA
D017122:Bacteriophage T4 / Q000378:metabolism; D052160:Benzothiazoles; D004265:DNA Helicases / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003959:Diamines; D005090:Exodeoxyribonucleases; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D009930:Organic Chemicals; D011485:Protein Binding; D011804:Quinolines; D053484:RecQ Helicases; D011994:Recombinant Proteins / Q000378:metabolism; D051716:Replication Protein A; D013329:Structure-Activity Relationship; D014764:Viral Proteins / Q000378:metabolism; D000071657:Werner Syndrome Helicase
D052160:Benzothiazoles; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D003959:Diamines; D005456:Fluorescent Dyes; D009930:Organic Chemicals; D011804:Quinolines; C497167:RPA1 protein, human; D011994:Recombinant Proteins; D051716:Replication Protein A; D014764:Viral Proteins; C060035:gp32 protein, Enterobacteria phage T4; C098022:SYBR Green I; D005090:Exodeoxyribonucleases; D004265:DNA Helicases; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase