INDUCTION OF NONBILAYER STRUCTURES IN DIACYLPHOSPHATIDYLCHOLINE MODELMEMBRANES BY TRANSMEMBRANE ALPHA-HELICAL PEPTIDES - IMPORTANCE OF HYDROPHOBIC MISMATCH AND PROPOSED ROLE OF TRYPTOPHANS

We have investigated the effect of several hydrophobic polypeptides on the phase behavior of diacylphosphatidylcholines with different acyl chain length, The polypeptides are uncharged and consist of a sequence with variable length of alternating leucine and alanine, flanked on b oth sides by two tryptophans, and with the N- and C-termini blocked. F irst it was demonstrated by circular dichroism measurements that these peptides adopt an ct-helical conformation with a transmembrane orient ation in bilayers of dimyristoylphosphatidylcholine. Subsequent P-31 N MR measurements showed that the peptides can affect lipid organization depending on the difference in hydrophobic length between the peptide and the lipid bilayer in the liquid-crystalline phase. When a 17 amin o acid residue long peptide (WALP17) was incorporated in a 1/10 molar ratio of peptide to lipid, a bilayer was maintained in saturated phosp holipids containing acyl chains of 12 and 14 C atoms, an isotropic pha se was formed at 16 C atoms, and an inverted hexagonal (H-II) phase at 18 and 20 C atoms. For a 19 amino acid residue long peptide (WALP19) similar changes in lipid phase behavior were observed, but at acyl cha in lengths of 2 C-atoms longer. Also in several cis-unsaturated phosph atidylcholine model membranes it was found that these peptides and a s horter analog (WALP16) induce the formation of nonbilayer structures a s a consequence of hydrophobic mismatch. It is proposed that this uniq ue ability of the peptides to induce nonbilayer structures in phosphat idylcholine model membranes is due to the presence of two tryptophans at both sides of the membrane/water interface, which prevent the pepti de from aggregating when the mismatch is increased. Comparison of the hydrophobic length of the bilayers with the length of the different pe ptides showed that it is the precise extent of mismatch that determine s whether the preferred lipid organization is a bilayer, isotropic pha se, or Hn phase. The peptide-containing bilayer and H!I phase were fur ther characterized after sucrose density gradient centrifugation of mi xtures of WALP16 and dioleoylphosphatidylcholine. P-31 NMR measurement s of the isolated fractions showed that a complete separation of both components was obtained. Chemical analysis of these fractions in sampl es with varying peptide concentration indicated that the H-II phase is highly enriched in peptide (peptide/lipid molar ratio of 1/6), while the maximum solubility of the peptide in the lipid bilayer is about 1/ 24 (peptide/lipid, molar). A molecular model of the peptide-induced HI I phase is presented that is consistent with the results obtained thus far.