A 39 amino acid fragment of the cell cycle regulator p21 is sufficient to bind PCNA and partially inhibit DNA replication in vivo.
The cell cycle regulator p21 interacts with and inhibits the DNA replication and repair factor proliferating cell nuclear antigen (PCNA). We have defined a 39 amino acid fragment of p21 which is sufficient to bind PCNA with high affinity (Kd 10-20 nM). This peptide can inhibit DNA replication in vitro and microinjection of a GST fusion protein containing this domain inhibited S phase in vivo. Despite its high affinity for PCNA, the free 39 amino acid peptide does not have a well-defined structure, as judged from circular dichroism and nuclear magnetic resonance measurements, suggesting an induced fit mechanism for the PCNA-p21 interaction. The association of the small peptide with PCNA was thermolabile, suggesting that portions of p21 adjoining the minimal region of contact stabilize the interaction. In addition, a domain containing 67 amino acids from the N-terminus of PCNA was defined as both necessary and sufficient for binding to p21.
Chen
J
J
Department of Pathology, Division of Molecular Oncology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Peters
R
R
Saha
P
P
Lee
P
P
Theodoras
A
A
Pagano
M
M
Wagner
G
G
Dutta
A
A
D000595:Amino Acid Sequence; D002478:Cells, Cultured; D050759:Cyclin-Dependent Kinase Inhibitor p21; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors; D016213:Cyclins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D004261:DNA Replication / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors; D005347:Fibroblasts; D005982:Glutathione Transferase / Q000235:genetics; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D018809:Proliferating Cell Nuclear Antigen / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D016196:S Phase / Q000187:drug effects; D017384:Sequence Deletion
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004791:Enzyme Inhibitors; D010446:Peptide Fragments; D018809:Proliferating Cell Nuclear Antigen; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D018844:Cyclin-Dependent Kinases