Purification of DNA polymerase delta as an essential simian virus 40 DNA replication factor

Melendy, T., Stillman, B. (1991) Purification of DNA polymerase delta as an essential simian virus 40 DNA replication factor. Journal of Biological Chemistry, 266 (3). pp. 1942-9. ISSN 0021-9258

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URL: http://www.ncbi.nlm.nih.gov/pubmed/1671044

Abstract

DNA replication from the SV40 origin can be reconstituted in vitro using purified SV40 large T antigen, cellular topoisomerases I and II, replication factor A (RF-A), proliferating cell nuclear antigen (PCNA), replication factor C (RF-C), and a phosphocellulose fraction (IIA) made from human cell extracts (S100). Fraction IIA contains all DNA polymerase activity required for replication in vitro in addition to other factors. A newly identified factor has been purified from fraction IIA. This factor is required for complete reconstitution of SV40 DNA replication and co-purifies with a PCNA-stimulated DNA polymerase activity. This DNA polymerase activity is sensitive to aphidicolin, but is not inhibited by butylanilinodeoxyadenosine triphosphate or by monoclonal antibodies which block synthesis by DNA polymerase alpha. The polymerase activity is synergistically stimulated by the combination of RF-A, PCNA, and RF-C in an ATP-dependent manner. Purified calf thymus polymerase delta can fully replace the purified factor in DNA replication assays. We conclude that this factor, required for reconstitution of SV40 DNA replication in vitro, corresponds to human DNA polymerase delta.

Item Type: Paper
Uncontrolled Keywords: Cell Fractionation DNA Polymerase II immunology DNA Polymerase III DNA Replication DNA-Directed DNA Polymerase antagonists & inhibitors immunology isolation purification metabolism Humans Hydrogen-Ion Concentration In Vitro Nuclear Proteins pharmacology Proliferating Cell Nuclear Antigen Simian virus 40 genetics Virus Replication
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > enzymes > DNA polymerase
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > DNA replication
CSHL Authors:
Communities: CSHL labs > Stillman lab
Highlight: Stillman, Bruce W.
Depositing User: CSHL Librarian
Date Deposited: 02 Mar 2012 19:33
Last Modified: 20 Jun 2017 20:10
Related URLs:
URI: http://repository.cshl.edu/id/eprint/25018

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