SKN-1 - EVIDENCE FOR A BIPARTITE RECOGNITION HELIX IN DNA-BINDING

Skn-1 is a maternally expressed transcription factor that specifies th e fate of certain blastomeres early in the development of Caenorhabdit is elegans. This transcription factor contains a basic region, but it binds to DNA as a monomer, Because other transcription factors contain ing basic regions bind as dimers, this finding implied that Skn repres ents a new DNA recognition motif, It has been proposed that the basic region helix of Skn is stabilized for binding by tertiary contacts to other parts of the protein, We have tested this proposal by carrying o ut circular dichroism (CD) and NMR experiments on the Skn domain and f ive truncated proteins, Our results have shown that the basic region o f Skn is unstructured in solution and does not contact other parts of the protein; like other basic region peptides, it folds into a helix o nly upon binding specifically to DNA. However, there is a stably folde d helical module in the Skn domain, and one of the helices in this mod ule terminates immediately before the start of the basic region. This pre-organized helix contains a surface rich in basic amino acids, and we propose that this helix contacts the DNA distal to the basic region proper, providing an extra long helical recognition surface which hel ps to stabilize monomeric binding, Homology between the Skn domain and several basic-region leucine zipper (bZIP) domains raises the possibi lity that the affinity and perhaps the specificity of DNA binding by b ZIP proteins can be modulated by incorporating a stably folded helical segment that contacts the DNA just below the basic region proper.