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Dimerization of CtIP, a BRCA1- and CtBP-interacting protein, is mediated by an N-terminal coiled-coil motif.

CtIP is a transcriptional co-regulator that binds a number of proteins involved in cell cycle control and cell development, such as CtBP (C terminus-binding protein), BRCA1 (breast cancer-associated protein-1), and LMO4 (LIM-only protein-4). The only recognizable structural motifs within CtIP are two putative coiled-coil domains located near the N and C termini of the protein. We now show that the N-terminal coiled coil (residues 45-160), but not the C-terminal coiled coil, mediates homodimerization of CtIP in mammalian 293T cells. The N-terminal coiled coil did not facilitate binding to LMO4 and BRCA1 proteins in these cells. A protease-resistant domain (residues 27-168) that minimally encompasses the putative N-terminal coiled coil was identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. This region is predicted to contain two smaller coiled-coil regions. The CtIP-(45-160) dimerization domain is helical and dimeric, indicating that the domain does form a coiled coil. The two smaller domains, CtIP-(45-92) and CtIP-(93-160), also formed dimers of lower binding affinity, but with less helical content than the longer peptide. The hydrodynamic radius of CtIP-(45-160) is the same as those of CtIP-(45-92) and CtIP-(93-160), implying that CtIP-(45-160) does not form a single long coiled coil, but a more compact structure involving homodimerization of the two smaller coiled coils, which fold back as a four-helix bundle or other compact structure. These results suggest a specific model for CtIP homodimerization via its N terminus and contribute to an improved understanding of how this protein might assemble other factors required for its role as a transcriptional corepressor.

Pubmed ID: 15084581

Authors

  • Dubin MJ
  • Stokes PH
  • Sum EY
  • Williams RS
  • Valova VA
  • Robinson PJ
  • Lindeman GJ
  • Glover JN
  • Visvader JE
  • Matthews JM

Journal

The Journal of biological chemistry

Publication Data

June 25, 2004

Associated Grants

None

Mesh Terms

  • Adaptor Proteins, Signal Transducing
  • Alcohol Oxidoreductases
  • Amino Acid Motifs
  • Amino Acid Sequence
  • BRCA1 Protein
  • Carrier Proteins
  • Cell Line
  • Chymotrypsin
  • Circular Dichroism
  • DNA-Binding Proteins
  • Dimerization
  • Escherichia coli
  • Homeodomain Proteins
  • Humans
  • LIM Domain Proteins
  • Models, Molecular
  • Molecular Sequence Data
  • Nuclear Proteins
  • Peptide Fragments
  • Phosphoproteins
  • Protein Structure, Tertiary
  • Recombinant Proteins
  • Retinoblastoma Protein
  • Transcription Factors
  • Transfection
  • Trypsin
  • Ultracentrifugation