Identification of DNA replication and cell cycle proteins that interact with PCNA

Gabriel Loor, Shan Jian Zhang, Peng Zhang, N. Lan Toomey, Marietta Y.W.T. Lee

Research output: Contribution to journalArticlepeer-review

74 Scopus citations

Abstract

The identity of DNA replication proteins and cell cycle regulatory proteins which can be found in complexes involving PCNA were investigated by the use of PCNA immobilized on Sepharose 4B. A column containing bovine serum albumin (BSA) bound to Sepharose was used as a control. Fetal calf thymus extracts were chromatographed on PCNA-Sepharose and BSA-Sepharose. The columns were washed and then eluted with 0.5 M KCl. The salt eluates were examined for the presence of both DNA replication proteins (pol α, δ, ε, PCNA, RFC, RFA, DNA ligase I, NDH II, Topo I and Topo II) and cell cycle proteins (Cyclins A, B1, D1, D2, D3, E, CDK2, CDK4, CDK5 and p21) by western blotting with specific antibodies. The DNA replication proteins which bound to PCNA-Sepharose included DNA polymerase δ and ε, PCNA, the 37 and 40 kDa subunits of RFC, the 70 kDa subunit of RPA, NDH II and topoisomerase I. No evidence for the binding of DNA polymerase α, DNA ligase I or topoisomerase II was obtained. Of the cell cycle proteins investigated, CDK2, CDK4 and CDK5 were bound. This study presents strong evidence that PCNA is a component of protein complexes containing DNA replication, repair and cell cycle regulatory proteins.

Original languageEnglish (US)
Pages (from-to)5041-5046
Number of pages6
JournalNucleic acids research
Volume25
Issue number24
DOIs
StatePublished - Dec 15 1997
Externally publishedYes

ASJC Scopus subject areas

  • Genetics

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