Cognate DNA stabilizes the tumor suppressor p53 and prevents misfolding and aggregation

Biochemistry. 2009 Jul 7;48(26):6126-35. doi: 10.1021/bi9003028.

Abstract

The tumor suppressor protein p53 is a nuclear protein that serves as an important transcription factor. The region responsible for sequence-specific DNA interaction is located in its core domain (p53C). Although full-length p53 binds to DNA as a tetramer, p53C binds as a monomer since it lacks the oligomerization domain. It has been previously demonstrated that two core domains have a dimerization interface and undergo conformational change when bound to DNA. Here we demonstrate that the interaction with a consensus DNA sequence provides the core domain of p53 with enhanced conformational stability at physiological salt concentrations (0.15 M). This stability could be either increased or abolished at low (0.01 M) or high (0.3 M) salt concentrations, respectively. In addition, interaction with the cognate sequence prevents aggregation of p53C into an amyloid-like structure, whereas binding to a nonconsensus DNA sequence has no effect on p53C stability, even at low ionic strength. Strikingly, sequence-specific DNA binding also resulted in a large stabilization of full-length p53, whereas nonspecific sequence binding led to no stabilization. The effects of cognate DNA could be mimicked by high concentrations of osmolytes such as glycerol, which implies that the stabilization is caused by the exclusion of water. Taken together, our results show an enhancement in protein stability driven by specific DNA recognition. When cognate DNA was added to misfolded protein obtained after a pressurization cycle, the original conformation was mostly recovered. Our results may aid the development of therapeutic approaches to prevent misfolded species of p53.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anilino Naphthalenesulfonates / chemistry
  • Consensus Sequence
  • DNA / metabolism
  • DNA / pharmacology*
  • Fluorescent Dyes / chemistry
  • Glycerol / chemistry
  • Humans
  • Hydrostatic Pressure
  • Light
  • Osmolar Concentration
  • Polydeoxyribonucleotides / chemistry
  • Polydeoxyribonucleotides / metabolism
  • Protein Binding / genetics
  • Protein Conformation / drug effects
  • Protein Denaturation
  • Protein Folding / drug effects*
  • Protein Stability / drug effects
  • Recombinant Proteins / chemistry
  • Scattering, Radiation
  • Spectrometry, Fluorescence
  • Tumor Suppressor Protein p53 / chemistry*
  • Tumor Suppressor Protein p53 / genetics
  • Tumor Suppressor Protein p53 / metabolism*
  • Water / chemistry

Substances

  • Anilino Naphthalenesulfonates
  • Fluorescent Dyes
  • Polydeoxyribonucleotides
  • Recombinant Proteins
  • Tumor Suppressor Protein p53
  • Water
  • poly(dC-dG)
  • 5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate)
  • DNA
  • Glycerol