title: Skn-1: evidence for a bipartite recognition helix in DNA binding.
abstract: Skn-1 is a maternally expressed transcription factor that specifies the fate of certain blastomeres early in the development of Caenorhabditis elegans. This transcription factor contains a basic region, but it binds to DNA as a monomer. Because other transcription factors containing basic regions bind as dimers, this finding implied that Skn represents 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 out circular dichroism (CD) and NMR experiments on the Skn domain and five truncated proteins. Our results have shown that the basic region of Skn is unstructured in solution and does not contact other parts of the protein; like other basic region peptides, it folds into a helix only upon binding specifically to DNA. However, there is a stably folded helical module in the Skn domain, and one of the helices in this module 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 helps to stabilize monomeric binding. Homology between the Skn domain and several basic-region leucine zipper (bZIP) domains raises the possibility that the affinity and perhaps the specificity of DNA binding by bZIP proteins can be modulated by incorporating a stably folded helical segment that contacts the DNA just below the basic region proper.
authors.affiliation: Department of Chemistry, Princeton University, Princeton, NJ 08544, USA.
authors.forename: S
authors.initials: S
authors.lastname: Pal
authors.forename: M C
authors.initials: MC
authors.lastname: Lo
authors.forename: D
authors.initials: D
authors.lastname: Schmidt
authors.forename: I
authors.initials: I
authors.lastname: Pelczer
authors.forename: S
authors.initials: S
authors.lastname: Thurber
authors.forename: S
authors.initials: S
authors.lastname: Walker
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D017173:Caenorhabditis elegans / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D017931:DNA Primers; D004268:DNA-Binding Proteins; D004926:Escherichia coli; D015801:Helminth Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016133:Polymerase Chain Reaction; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D013816:Thermodynamics; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism
chemical_list: D029742:Caenorhabditis elegans Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D015801:Helminth Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; C073702:skn-1 protein, C elegans; D004247:DNA