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PMID: 3035543 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Initiation of simian virus 40 DNA replication in vitro: large-tumor-antigen- and origin-dependent unwinding of the template.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 84 ·No. 11 ·1987-06-00 ·Pages 3643-7

Wold MS, Li JJ, Kelly TJ

Abstract

Analysis of the kinetics of simian virus 40 (SV40) DNA replication in vitro demonstrated the existence of a slow presynthesis reaction that occurs prior to onset of extensive chain elongation and is dependent on a subset of the cellular proteins required for the complete replication reaction. When the presynthesis reaction is carried out in the presence of topoisomerase I, it is possible to detect extensive unwinding of the template DNA. This unwinding reaction is specific for templates that contain the wild-type SV40 origin of DNA replication and requires SV40 large tumor antigen (T antigen), ATP, and a protein fraction derived from HeLa cells. The required cellular protein may be a eukaryotic single-stranded-DNA-binding protein (SSB), since unwinding of the template is also observed when Escherichia coli SSB is substituted for the HeLa protein fraction. These observations suggest that during the initial stages of SV40 DNA replication, T antigen binds specifically to the viral origin and locally unwinds the DNA. This origin-dependent unwinding reaction is presumably a prerequisite for subsequent priming and elongation steps.

MeSH Terms
Antigens, Polyomavirus Transforming Antigens, Viral, Tumor/metabolism Cell-Free System DNA Replication DNA, Viral/genetics,ultrastructure Genetic Vectors Kinetics Oncogene Proteins, Viral/metabolism Protein Kinases/metabolism Simian virus 40/enzymology,genetics Templates, Genetic
Chemicals
Antigens, Polyomavirus Transforming Antigens, Viral, Tumor DNA, Viral Oncogene Proteins, Viral Protein Kinases
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Wold M S
Li J J
Kelly T J
References (28)
28 references, click to expand
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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.84.11.3643
Published
1987-06-00
Pages
3643-7
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC304931
Subset
IM
Grants
NCI NIH HHS · CA09139 · United States
NIGMS NIH HHS · GM07309 · United States
External Links
PubMed source DOI: 10.1073/pnas.84.11.3643 PMC Full Text View journal details
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