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PMID: 2548199 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Template supercoiling during ATP-dependent DNA helix tracking: studies with simian virus 40 large tumor antigen.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 86 ·No. 16 ·1989-08-00 ·Pages 6121-5

Yang L, Jessee CB, Lau K, Zhang H, Liu LF

Abstract

Incubation of topologically relaxed plasmid DNA with simian virus 40 (SV40) large tumor antigen (T antigen), ATP, and eubacterial DNA topoisomerase I resulted in the formation of highly positively supercoiled DNA. Eukaryotic DNA topoisomerase I could not substitute for eubacterial DNA topoisomerase 1 in this reaction. Furthermore, the addition of eukaryotic topoisomerase I to a preincubated reaction mixture containing both T antigen and eubacterial topoisomerase I caused rapid relaxation of the positively supercoiled DNA. These results suggest that SV40 T antigen can introduce topoisomerase-relaxable supercoils into DNA in a reaction coupled to ATP hydrolysis. We interpret the observed T antigen supercoiling reaction in terms of a recently proposed twin-supercoiled-domain model that describes the mechanics of DNA helix-tracking processes. According to this model positive and negative supercoils are generated ahead of and behind the moving SV40 T antigen, respectively. The preferential relaxation of negative supercoils by eubacterial DNA topoisomerase I explains the accumulation of positive supercoils in the DNA template. The supercoiling assay using DNA conformation-specific eubacterial DNA topoisomerase I may be of general use for the detection of ATP-dependent DNA helix-tracking proteins.

MeSH Terms
Adenosine Triphosphate/metabolism Antigens, Polyomavirus Transforming/genetics,isolation & purification Cell Line DNA Topoisomerases, Type I/metabolism DNA, Superhelical/genetics,isolation & purification Electrophoresis, Gel, Two-Dimensional Escherichia coli/enzymology Humans Nucleic Acid Conformation Plasmids Polyethylene Glycols/pharmacology Simian virus 40/genetics Templates, Genetic
Chemicals
Antigens, Polyomavirus Transforming DNA, Superhelical Polyethylene Glycols Adenosine Triphosphate DNA Topoisomerases, Type I
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Yang L
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205.
Jessee C B
Lau K
Zhang H
Liu L F
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.86.16.6121
Published
1989-08-00
Pages
6121-5
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC297787
Subset
IM
Grants
NIGMS NIH HHS · GM27731 · United States
External Links
PubMed source DOI: 10.1073/pnas.86.16.6121 PMC Full Text View journal details
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