title: Multiple amino acids determine the DNA binding specificity of the Msx-1 homeodomain.
abstract: This study investigates the sequence features that contribute to the differential DNA binding properties of two divergent homeodomains, Msx-1 and HoxA3. We show that these homeodomains have overlapping, but nonidentical, DNA binding site preferences. We defined the amino acid residues that contribute to the observed differences in DNA binding specificity by producing a series of mutated polypeptides in which selected residues in Msx-1 were replaced with the corresponding ones in HoxA3. These analyses show that the DNA binding specificity of Msx-1 versus HoxA3 results from the cumulative action of multiple residues in all segments of the homeodomain (i.e., the N-terminal arm and helices I, II, and III). Therefore, substitutions of residues in both helix III and the N-terminal arm (but not in either segment alone) produced an Msx-1 polypeptide whose binding site preference was indistinguishable from that of HoxA3. Residues in helices I and II also influence DNA binding activity; these oppositely charged residues (e.g., lysine 19 and glutamate 30) may mediate ionic interactions between helices I and II which stabilize DNA binding by Msx-1. These findings demonstrate a critical interplay between residues in each homeodomain segment for appropriate conformation of the protein-DNA complex.
authors.affiliation: Center for Advanced Biotechnology and Medicine, Piscataway, New Jersey 08854-5638, USA.
authors.forename: V E
authors.initials: VE
authors.lastname: Isaac
authors.forename: P
authors.initials: P
authors.lastname: Sciavolino
authors.forename: C
authors.initials: C
authors.lastname: Abate
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000737:chemistry / Q000378:metabolism*; D051957:MSX1 Transcription Factor; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D014157:Transcription Factors
chemical_list: D000596:Amino Acids; D018398:Homeodomain Proteins; D051957:MSX1 Transcription Factor; D014157:Transcription Factors; D004247:DNA