TOPOGRAPHY OF THE MEMBRANE-BINDING DOMAIN OF CYTOCHROME-B5 IN LIPIDS BY FOURIER-TRANSFORM INFRARED-SPECTROSCOPY

Fourier-transform infrared spectroscopy was used to examine the secondary structure of the membrane-binding domain (nonpolar peptide) of rabbit liver cytochrome b5 in D2O and in the presence of phospholipids and deoxycholate. In all situations, the predominant structure was α helix, but an examination of the components of the amide I band in the spectrum of the nonpolar peptide showed that the major peak was shifted from 1655 cm−1 in the lipids to 1650 cm−1 in deoxycholate. This shift to lower frequency, together with a decrease in intensity of the amide II band, is indicative of N-H to N-D exchange of the peptide backbone. A semiquantitative analysis indicated that the α helix of the peptide is over 95% exchanged in the presence of deoxycholate but is only 10% exchanged in the presence of lipid. These data suggest that the membrane-inserted portion of the peptide is α helical and is largely protected from N-H to N-D exchange by the bilayer. We suggest that this technique appears to provide a general method for determining the type of secondary structure involved in membrane interaction and the percentage of this structure which is involved in the interaction. © 1990, American Chemical Society. All rights reserved.