Studies on the activator 1 protein complex, an accessory factor for proliferating cell nuclear antigen-dependent DNA polymerase δ

Suk-Hee Lee, Ann D. Kwong, Zhen Qiang Pan, Jerard Hurwitz

Research output: Contribution to journalArticle

192 Citations (Scopus)

Abstract

Activator 1 (A1) is a multiprotein complex which is essential for proliferating cell nuclear antigen (PCNA)-dependent DNA polymerase δ (pol δ) activity and efficient in vitro DNA synthesis in the SV40 dipolymerase replication system. In this report, we describe the isolation of A1 from HeLa cytosolic extracts. A1 stimulated pol δ activity in singly primed øX174 DNA or (dA)4500·oligo(dT)12-18 in reactions containing PCNA, single-stranded DNA binding protein (SSB), and ATP. Using this assay, A1 has been extensively purified. Purified preparations contained five discrete subunits of 145, 40, 38, 37, and 36.5 kDa. ATP hydrolysis to ADP and Pi is essential for A1-dependent pol δ activity, and we have shown that A1 contains an intrinsic ATPase which is stimulated by DNA. The DNA-dependent hydrolysis of ATP can be stimulated by PCNA and further activated by PCNA plus the human single-stranded DNA binding protein. These stimulatory effects were observed with (dA)4500· oligo(dT)12-18, but were not detected with each polydeoxynucleotide alone. Furthermore, A1 formed a complex with (dA)4500·oligo(dT)12-18 which could be measured by nitrocellulose binding. No complex with (dA)4500 or oligo(dT)12-18 alone was detected by this procedure. Data are also presented which indicate that A1, in conjunction with PCNA, functions as a primer- recognition factor for pol δ, increasing its ability to utilize low levels of primer ends, but it does not increase the size of the DNA products. A1 also markedly reduced the amount of PCNA required for pol δ activity on a multiply primed DNA suggesting that PCNA interacts with A1 at the primer end. These multiple effects of A1 closely resemble the properties of the multisubunit protein RF-C described by Tsurimoto and Stillman (Tsurimoto, T., and Stillman, B. (1990) Proc. Natl. Acad. Sci. U. S. A. 87, 1023-1027).

Original languageEnglish (US)
Pages (from-to)594-602
Number of pages9
JournalJournal of Biological Chemistry
Volume266
Issue number1
StatePublished - Jan 5 1991
Externally publishedYes

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Replication Protein C
Proliferating Cell Nuclear Antigen
Accessories
DNA-Directed DNA Polymerase
DNA
Adenosine Triphosphate
DNA-Binding Proteins
Hydrolysis
Multiprotein Complexes
Collodion
Protein C
Adenosine Diphosphate
Adenosine Triphosphatases
Assays

ASJC Scopus subject areas

  • Biochemistry

Cite this

Studies on the activator 1 protein complex, an accessory factor for proliferating cell nuclear antigen-dependent DNA polymerase δ. / Lee, Suk-Hee; Kwong, Ann D.; Pan, Zhen Qiang; Hurwitz, Jerard.

In: Journal of Biological Chemistry, Vol. 266, No. 1, 05.01.1991, p. 594-602.

Research output: Contribution to journalArticle

Lee, Suk-Hee ; Kwong, Ann D. ; Pan, Zhen Qiang ; Hurwitz, Jerard. / Studies on the activator 1 protein complex, an accessory factor for proliferating cell nuclear antigen-dependent DNA polymerase δ. In: Journal of Biological Chemistry. 1991 ; Vol. 266, No. 1. pp. 594-602.
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