Nucleic Acids Research, Vol 27, Issue 17 3557-3566, Copyright © 1999 by Oxford University Press
DK Orren, RMJr, JO Nehlin, A Machwe, MD Gray and VA Bohr
Mutations in the WRN gene result in Werner syndrome, an autosomal recessive
disease in which many characteristics of aging are accelerated. A probable
role in some aspect of DNA metabolism is suggested by the primary sequence
of the WRN gene product. A recombinant His-tagged WRN protein (WRNp) was
overproduced in insect cells using the baculovirus system and purified to
near homogeneity by several chromatographic steps. This purification scheme
removes both nuclease and topoisomerase contaminants that persist following
a single Ni(2+)affinity chromatography step and allows for unambiguous
interpretation of WRNp enzymatic activities on DNA substrates. Purified
WRNp has DNA-dependent ATPase and helicase activities consistent with its
homology to the RecQ subfamily of proteins. The protein also binds with
higher affinity to single-stranded DNA than to double-stranded DNA.
However, WRNp has no higher affinity for various types of DNA damage,
including adducts formed during 4NQO treatment, than for undamaged DNA. Our
results confirm that WRNp has a role in DNA metabolism, although this role
does not appear to be the specific recognition of damage in DNA.
ARTICLES
Enzymatic and DNA binding properties of purified WRN protein: high affinity binding to single-stranded DNA but not to DNA damage induced by 4NQO
Laboratory of Molecular Genetics, National Institute on Aging, National Institutes of Health, Baltimore, MD 21224, USA.
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