Control of the transmembrane orientation and interhelical interactions within membranes by hydrophobic helix length

We examined the effect of the length of the hydrophobic core of Lys-flanked poly(Leu) peptides on their behavior when inserted into model membranes. P eptide structure and membrane location were assessed by the fluorescence em ission lambda(max) of a Trp residue in the center of the peptide sequence, the quenching of Trp fluorescence by nitroxide-labeled lipids (parallax ana lysis), and circular dichroism. Peptides in which the hydrophobic core vari ed in length from 11 to 23 residues were found to be largely alpha-helical when inserted into the bilayer. In dioleoylphosphatidylcholine (diC(18:1)PC ) bilayers, a peptide with a 19-residue hydrophobic core exhibited highly b lue-shifted fluorescence, an indication of Trp location in a nonpolar envir onment, and quenching localized the Trp to the bilayer center, an indicatio n of transmembrane structure. A peptide with an 11-residue hydrophobic core exhibited emission that was red-shifted, suggesting a more polar Trp envir onment, and quenching showed the Trp was significantly displaced from the b ilayer center, indicating that this peptide formed a nontransmembranous str ucture. A peptide with a 23-residue hydrophobic core gave somewhat red-shif ted fluorescence, but quenching demonstrated the Trp was still close to the bilayer center, consistent with a transmembrane structure. Analogous behav ior was observed when the behavior of individual peptides was examined in m odel membranes with various bilayer widths. Other experiments demonstrated that in diC(18:1)PC bilayers the dilution of the membrane concentration of the peptide with a 23-residue hydrophobic core resulted in a blue shift of fluorescence, suggesting the red-shifted fluorescence at higher peptide con centrations was due to helix oligomerization. The intermolecular self-quenc hing of rhodamine observed when the peptide was rhodamine-labeled, and the concentration dependence of self-quenching, supported this conclusion. Thes e studies indicate that the mismatch between helix length and bilayer width can control membrane location, orientation, and helix-helix interactions, and thus may mismatch control both membrane protein folding and the interac tions between membrane proteins.