title: Orientation in lipid bilayers of a synthetic peptide representing the C-terminus of the A1 domain of shiga toxin. A polarized ATR-FTIR study.
abstract: The interaction of a synthetic peptide representing the C-terminal 27 amino acids of the A1 domain of Shiga toxin (residues 220-246) with acidic phospholipid model membranes was characterized by FTIR spectroscopy. This peptide resembles a signal sequence and may mediate the translocation of the catalytic A1 chain of Shiga toxin to the cytoplasm following its retrograde transport to the lumenal compartment of the endoplasmic reticulum (ER). At pH 7 and 5, the peptide underwent a conformational change from random coil to alpha-helix upon addition of negatively charged phospholipids. Examination of the amide II band in the spectrum of the complex at pH 7 and pH 5 showed that in both cases, the N-H groups in the peptide backbone are largely protected from H/D exchange. Using polarized attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) measurements, the orientation of the alpha-helical portion of the peptide was found to be almost perpendicular with respect to the membrane plane at pH 7. However, at pH 5.0-5.4, the alpha-helix axis was preferentially oriented parallel to the membrane plane. The results suggest that at the neutral pH of the ER lumen, the peptide may insert into the membrane, while at the lower pH levels present in earlier endocytic compartments, the peptide would be less likely to traverse the bilayer. In summary, this putative signal peptide may not be able to cause a significant translocation of the A1 domain of Shiga toxin to the cytosol until it reaches the neutral pH of the ER compartment.
authors.affiliation: The Research Institute, The Hospital for Sick Children, Toronto, Canada M5G 1X8.
authors.forename: A
authors.initials: A
authors.lastname: Menikh
authors.forename: M T
authors.initials: MT
authors.lastname: Saleh
authors.forename: J
authors.initials: J
authors.lastname: Gariépy
authors.forename: J M
authors.initials: JM
authors.lastname: Boggs
mesh_terms: D000577:Amides / Q000737:chemistry; D000595:Amino Acid Sequence; D001427:Bacterial Toxins / Q000737:chemistry* / Q000378:metabolism; D017666:Deuterium Oxide; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism; D010710:Phosphates / Q000737:chemistry; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D021382:Protein Sorting Signals / Q000737:chemistry / Q000378:metabolism; D017433:Protein Structure, Secondary; D022603:Shiga Toxins; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D000577:Amides; D001427:Bacterial Toxins; D008051:Lipid Bilayers; D010446:Peptide Fragments; D010710:Phosphates; D010743:Phospholipids; D021382:Protein Sorting Signals; D022603:Shiga Toxins; D017666:Deuterium Oxide