A β-sheet peptide inhibitor of E47 dimerization and DNA binding Original Research Article

Abstract 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 β-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 β-sheet secondary structure upon the highly a-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 β-sheet induction in an a-helical protein. Future experiments will probe the structural determinants of the β-sheet secondary structure in helix II and investigate the generality of the dissociative strategy in other transcription factor families.