A beta-sheet peptide inhibitor of E47 dimerization and DNA binding.
BACKGROUND
Many transcription factors are active only in their dimeric form, including the basic-helix-loop-helix (bHLH) family of transcription factors. The disruption of the dimer therefore presents a means of inhibiting the biological functions of such transcription factors. E47 is a homodimeric bHLH transcription factor with a four-helix bundle dimerization interface. Here, we investigate the concept of dimerization inhibition using peptides derived from the dimerization domain of E47.


RESULTS
We have synthesized several peptides corresponding to the E47 dimerization interface that inhibit E47 DNA-binding activity with IC50 values in the range of 3.6-120 mM. Interestingly, helix II; a peptide corresponding to the carboxy-terminal helix of the E47 dimerization interface, adopted a beta-sheet structure in solution, as shown using circular dichroism (CD), and inhibited the binding of E47 to DNA at equimolar concentrations. Size-exclusion chromatography, analytical ultracentrifugation and cross-linking experiments verified that this peptide prevented E47 dimerization. Furthermore, CD experiments provided evidence that helix II could induce a beta-sheet secondary structure upon the highly alpha-helical E47 bHLH domain.


CONCLUSIONS
This study is the first demonstration of dissociative inhibition in the bHLH class of transcription factors and also provides an example of beta-sheet induction in an alpha-helical protein. Future experiments will prove the structural determinants of the beta-sheet secondary structure in helix II and investigate the generality of the dissociative strategy in other transcription factor families.
Ghosh
I
I
Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA.
Chmielewski
J
J
D000595:Amino Acid Sequence; D001665:Binding Sites; D002850:Chromatography, Gel; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D018257:Helix-Loop-Helix Motifs; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology*; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors / Q000378:metabolism*; D014461:Ultracentrifugation
D004268:DNA-Binding Proteins; D010446:Peptide Fragments; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D014157:Transcription Factors; D004247:DNA