Horst Vogel
The secondary structure of alamethicin in lipid membranes below and above the lipid phase transition temp. Tt is detd. by Raman spectroscopy and CD measurements. In both cases structural data are obtained by fitting the exptl. spectra by a superposition of the spectra of 15 ref. proteins of known 3-dimensional structure. According to the Raman expts., in a lipid bilayer above Tt, alamethicin is helical from residue 1 to 12, whereas below Tt the helix extends from residue 1 to 16. The remaining C-terminal part is nonhelical up to the end residue 20 both above and below Tt. A considerably lower helix content is derived from CD, namely, 38% and 46% above and below Tt, resp., in agreement with several reported values for CD in the literature. It is shown that the commonly used set of CD spectra of water-sol. ref. proteins is unsuitable to describe the CD spectra of alamethicin correctly. Therefore, the secondary structure of alamethicin as derived from CD measurement is, at the present state of anal., unreliable. In contrast to the case of alamethicin, the CD spectra of melittin in lipid membranes are correctly described by the ref. protein spectra. The helix content of melittin is detd. thereby to be 72% in lipid membranes above Tt and 75% below Tt. The data are in accord with a structure where the hydrophobic part of melittin adopts a bent helix as detd. recently by Raman spectroscopy. The orientation order parameters of the helical parts of alamethicin and of melittin in a lipid membrane are deduced from the difference between a corresponding CD spectrum of a polypeptide in planar multibilayer and that in lipid vesicles. The presented method for detg. helix order parameters is new and may be generally applicable to other membrane proteins. The orientation of the helical part of both polypeptides depends on the phys. state of the lipid bilayer at maximal membrane hydration and in the ordered, lipid state furthermore on the degree of membrane hydration. Under conditions where alamethicin and melittin are incorporated in an aggregated form in a fluid lipid membrane at maximal water content, the helical segments are oriented preferentially parallel to the membrane normal. Cooling such lipid membranes to a temp. below Tt changes the orientation of the helical part of alamethicin as well as melittin toward the membrane plane. On the contrary, in dried planar membranes at 2% water content, both polypeptide mols. are oriented with their helical parts parallel to the membrane normal, similar to the case of fluid lipid membranes. [on SciFinder (R)]
1987
