cdc2 family kinases phosphorylate a human cell DNA replication factor, RPA, and activate DNA replication

Dutta, A., Stillman, B. (June 1992) cdc2 family kinases phosphorylate a human cell DNA replication factor, RPA, and activate DNA replication. Embo J, 11 (6). pp. 2189-99. ISSN 0261-4189 (Print)



RPA is a single-stranded DNA binding protein complex purified from human cells and is essential for the initiation and elongation stages of SV40 DNA replication in vitro. In both human and yeast cells, the 34 kDa polypeptide subunit of RPA is phosphorylated in the S and G2 phases of the cell cycle and not in G1. One of the major RPA kinases present in extracts of human cells was purified and shown to be the cyclin B-cdc2 complex. This purified kinase, and a closely related cyclin A associated cdc2-like kinase, phosphorylated RPA p34 on a subset of the chymotryptic peptides that were phosphorylated in vivo at the G1-S transition. Two serines near the N-terminus of RPA p34 were identified as possible sites of phosphorylation by cdc2 kinase. These same serines were necessary for RPA phosphorylation in vivo. The purified cdc2 kinase stimulated SV40 DNA replication in vitro when added to G1 cell extracts. The kinase also stimulated unwinding at the origin of replication, one of the earliest steps in DNA replication requiring RPA, but only in the presence of an additional factor present in G1 cell extracts. Thus, one or more members of the cyclin-cdc2 kinase family may be required for the initiation and maintenance of S phase, in part due to their ability to phosphorylate and activate a cellular DNA replication factor, RPA.

Item Type: Paper
Uncontrolled Keywords: Adenovirus Early Proteins Amino Acid Sequence CDC2 Protein Kinase metabolism Cell Cycle physiology Cell Line Transformed Cyclins metabolism DNA Replication DNA-Binding Proteins genetics/isolation & purification metabolism G1 Phase physiology Hela Cells Humans Kinetics Molecular Sequence Data Mutagenesis Site-Directed Oncogene Proteins Viral genetics Peptide Mapping Phosphopeptides isolation & purification Phosphorylation Replication Factor A S Phase/physiology Simian virus 40 genetics
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > DNA replication
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein expression > phosphorylation
CSHL Authors:
Communities: CSHL labs > Stillman lab
Highlight: Stillman, Bruce W.
Depositing User: CSHL Librarian
Date: June 1992
Date Deposited: 06 Mar 2012 22:01
Last Modified: 20 Jun 2017 20:02
PMCID: PMC556686

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