NDLI: The SF1 helicase encoded by the archaeal plasmid pTN2 of Thermococcus nautili

Content Provider	Springer Nature Link
Author	Béguin, Pierre Baron, Bru Gill, Sukhvinder Charpin, Nicole Forterre, Patrick
Copyright Year	2014
Abstract	We expressed, purified, and characterized the helicase encoded by ORF1 of the Thermococcus nautili pTN2 plasmid (Soler et al. Nucl Acids Res 38, 5088–5104, 2010). The enzyme, which belongs to the SF1 family of helicases, possesses NTPase activity, with a strong preference for ATP and GTP as compared to CTP and TTP; dATP was also a substrate. Triphosphatase activity was strongly stimulated by single-stranded DNA and, to a lesser extent, by double-stranded DNA. Unwinding of duplexes comprising a fluorescent oligonucleotide was monitored by fluorescence polarization spectroscopy and by polyacrylamide gel electrophoresis. As observed for enzymes of the same family, pTN2 helicase displays a strong preference for duplexes comprising a 3′ single-stranded extension and proceeds from the 3′ to the 5′ end of the loading strand. Under the conditions of the in vitro assay, pTN2 helicase did not appear to be recycled, but stayed bound to single-stranded DNA, which explains why high concentrations of enzyme are required to unwind long stretches of duplex DNA. The helicase enhances the synthesis of double-stranded DNA by pTN2 primase and by T. nautili PolB polymerase primed by pTN2 primase but it did not enhance synthesis by Taq DNA polymerase.
Starting Page	779
Ending Page	787
Page Count	9
File Format	PDF
ISSN	14310651
Journal	Extremophiles
Volume Number	18
Issue Number	4
e-ISSN	14334909
Language	English
Publisher	Springer Japan
Publisher Date	2014-06-03
Publisher Place	Tokyo
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Helicase Archaeal DNA replication Thermococcus nautili Microbiology Biotechnology Biochemistry Microbial Ecology
Content Type	Text
Resource Type	Article
Subject	Medicine Molecular Medicine Microbiology

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