Conformational analysis by NMR and distance geometry techniques of a peptide mimetic of the third helix of the Antennapedia homeodomain.
The Antennapedia homeodomain structure consists of four helices. The helices II and III are connected by a tripeptide that forms a turn, and constitute the well-known helix-turn-helix motif. The recognition helix penetrates the DNA major groove, gives specific protein-DNA contacts and forms direct, or water-mediated, intermolecular hydrogen bonds. It was suggested that helix III (and perhaps also helix IV) might represent the recognition helix of Antennapedia homeodomain, which makes contact with the surface of the major groove of the DNA. In an attempt to clarify the helix III capabilities of assuming an helical conformation when separated from the rest of the protein, we carried out the structural determination of the recognition helix III in different solvent media. The conformational study of fragments 42-53, where residues W48 and F49, not involved in the protein-DNA interaction, were substituted by two alanines, was conducted in sodium dodecyl sulfate (SDS), trifluoroethanol (TFE) and TFE/water, using circular dichroism, nuclear magnetic resonance (NMR) and distance geometry (DG) techniques. The fragment assumes a well-defined secondary structure in TFE and in TFE/water (90/10, v/v) with an alpha-helix encompassing residues 4-9, while in TFE/water (70/30, v/v) a less regular structure was found. The DG results in the micellar system evidence the presence of a distorted alpha-helical conformation involving residues 4-8. Our results reveal that the isolated Antennapedia recognition helix III tend to preserve in solution the alpha-helical conformation even if separated from the rest of the molecule.
Saviano
M
M
Institute of Biostructures and Bioimaging, CNR, via Mezzocannone 16, I-80134 Napoli, Italy.
Isernia
C
C
Bassarello
C
C
Di Lello
P
P
Galdiero
S
S
Mierke
D F
DF
Benedetti
E
E
Pedone
C
C
D000818:Animals; D051943:Antennapedia Homeodomain Protein; D018398:Homeodomain Proteins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D009687:Nuclear Proteins / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry*; D017433:Protein Structure, Secondary; D012997:Solvents / Q000737:chemistry; D014157:Transcription Factors / Q000737:chemistry*; D014270:Trifluoroethanol / Q000737:chemistry
D051943:Antennapedia Homeodomain Protein; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; D010455:Peptides; D012997:Solvents; D014157:Transcription Factors; D014270:Trifluoroethanol