Human-Saccharomyces cerevisiae proliferating cell nuclear antigen hybrids - Oligomeric structure and functional characterization using in vitro DNA replication

Ola, A., Waga, S., Ellison, V., Stillman, B., McGurk, M., Leigh, I. M., Waseem, N. H., Waseem, A. (2001) Human-Saccharomyces cerevisiae proliferating cell nuclear antigen hybrids - Oligomeric structure and functional characterization using in vitro DNA replication. Journal of Biological Chemistry, 276 (13). pp. 10168-10177. ISSN 0021-9258

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URL: http://www.ncbi.nlm.nih.gov/pubmed/11094057
DOI: 10.1074/jbc.M008929200

Abstract

The proliferating cell nuclear antigen (PCNA) is a highly conserved protein required for the assembly of the DNA polymerase delta (pol delta) holoenzyme, Because PCNAs from Saccharomyces cerevisiae and human do not complement each other using in vitro or in vivo assays, hybrids of the two proteins would help identify region(s) involved in the assembly of the pol S holoenzyme, Two mutants of human PCNA, HU1 (D21E) and HU3 (D120E), and six hybrids of human and S, cerevisiae PCNA, HC1, HC5, CH2, CH3, CH4, and CH5, were prepared by swapping corresponding regions between the two proteins, In solution, all PCNA assembled into trimers, albeit to different extents, These PCNA variants were tested for stimulation of pol delta and in vitro replication of M13 and SV40 DNA as well as to stimulate the ATPase activity of replication factor C (RF-C), Our data suggest that in addition to the interdomain connecting loop and C terminus, an additional site in the N terminus is required for pol delta interaction. PCNA mutants and hybrids that stimulated pol delta and RF-C were deficient in M13 and SV40 DNA replication assays, indicating that PCNA-induced pol delta stimulation and RF-C-mediated loading are not sufficient for coordinated DNA synthesis at a replication fork.

Item Type: Paper
Uncontrolled Keywords: SIMIAN VIRUS-40 DNA SV-40 DNA Simian Virus 40 LAGGING STRAND SYNTHESIS lagging strand synthesis DELTA-AUXILIARY PROTEIN delta-auxiliary protein C-TERMINAL REGION C-terminal region C terminal region POLYMERASE-DELTA Polyphase_Delta HUMAN PCNA Human pCNA ACCESSORY PROTEINS accessory proteins TRIMERIC STRUCTURE trimeric structure MISMATCH REPAIR mismatch repair MULTIPLE STAGES multiple stages
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > DNA replication
organism description > yeast > Saccharomyces
CSHL Authors:
Communities: CSHL labs > Stillman lab
Highlight: Stillman, Bruce W.
Depositing User: CSHL Librarian
Date Deposited: 02 Mar 2012 19:16
Last Modified: 20 Jun 2017 18:49
Related URLs:
URI: http://repository.cshl.edu/id/eprint/25034

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