A 39 amino acid fragment of the cell cycle regulator p21 is sufficient to bind PCNA and partially inhibit DNA replication in vivo

Nucleic Acids Res. 1996 May 1;24(9):1727-33. doi: 10.1093/nar/24.9.1727.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Cells, Cultured
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclin-Dependent Kinases / antagonists & inhibitors
  • Cyclins / chemistry*
  • Cyclins / metabolism
  • Cyclins / pharmacology
  • DNA Replication / drug effects
  • DNA Replication / physiology*
  • Enzyme Inhibitors
  • Fibroblasts
  • Glutathione Transferase / genetics
  • Humans
  • Kinetics
  • Molecular Sequence Data
  • Peptide Fragments / chemical synthesis
  • Peptide Fragments / chemistry*
  • Peptide Fragments / metabolism
  • Peptide Fragments / pharmacology
  • Proliferating Cell Nuclear Antigen / chemistry
  • Proliferating Cell Nuclear Antigen / genetics
  • Proliferating Cell Nuclear Antigen / metabolism*
  • Protein Binding
  • Protein Structure, Secondary
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / metabolism
  • Recombinant Fusion Proteins / pharmacology
  • S Phase / drug effects
  • Sequence Deletion

Substances

  • CDKN1A protein, human
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclins
  • Enzyme Inhibitors
  • Peptide Fragments
  • Proliferating Cell Nuclear Antigen
  • Recombinant Fusion Proteins
  • Glutathione Transferase
  • Cyclin-Dependent Kinases