ORIENTATION IN LIPID BILAYERS OF A SYNTHETIC PEPTIDE REPRESENTING THEC-TERMINUS OF THE A(1) DOMAIN OF SHIGA TOXIN - A POLARIZED ATR-FTIR STUDY

The interaction of a synthetic peptide representing the C-terminal 27 amino acids of the A(1) domain of Shiga toxin (residues 220-246) with acidic phospholipid model membranes was characterized by FTIR spectros copy. This peptide resembles a signal sequence and may mediate the tra nslocation of the catalytic Al chain of Shiga toxin to the cytoplasm f ollowing its retrograde transport to the lumenal compartment of the en doplasmic reticulum (ER). At pH 7 and 5, the peptide underwent a confo rmational change from random coil to a-helix upon addition of negative ly charged phospholipids. Examination of the amide II band in the spec trum 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 exchan ge. Using polarized attenuated total reflectance Fourier transform inf rared spectroscopy (ATR-FTIR) measurements, the orientation of the or- helical portion of the peptide was found to be almost perpendicular wi th 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 pr esent in earlier endocytic compartments, the peptide would be less lik ely to traverse the bilayer. In summary, this putative signal peptide may not be able to cause a significant translocation of the A(1) domai n of Shiga toxin to the cytosol until it reaches the neutral pH of the ER compartment.