Partitioning of lipidated peptide sequences into liquid-ordered lipid domains in model and biological membranes.
We have used a fluorescence assay and detergent fractionation to examine the partitioning of different fluorescent lipidated peptides, with sequences and lipid substituents matching those found in various classes of lipidated cellular proteins, into liquid-ordered (raft-like) domains in lipid bilayers. Peptides incorporating isoprenyl groups, or multiple unsaturated acyl chains, show negligible affinity for liquid-ordered domains in mixed-phase liquid-ordered/liquid-disordered (l(o)/l(d)) bilayers composed of dipalmitoylphosphatidylcholine, a spin-labeled unsaturated phosphatidylcholine, and cholesterol. By contrast, peptides incorporating multiple S- and/or N-acyl chains, or a cholesterol residue plus an N-terminal palmitoyl chain, show significant partitioning into liquid-ordered domains under the same conditions. Interestingly, the affinity of a lipidated peptide for l(o) domains can be strongly influenced, not only by the structures of the lipid substituents but also by the nature and the positions of their attachment to the peptide chain. These results are well correlated with those obtained from parallel assays based on low-temperature detergent fractionation. Using the latter approach, we further demonstrate that a truly minimal l(o) domain partitioning motif [myristoylGlyCys(palmitoyl)-] can mediate efficient incorporation into the "raft" fraction of COS-7 cell membranes.
Wang
T Y
TY
Department of Biochemistry, McGill University, Montréal, Québec, Canada H3G 1Y6.
Leventis
R
R
Silvius
J R
JR
D015060:1,2-Dipalmitoylphosphatidylcholine / Q000737:chemistry; D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000737:chemistry*; D002784:Cholesterol / Q000737:chemistry; D003902:Detergents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D049452:Green Fluorescent Proteins; D008051:Lipid Bilayers / Q000737:chemistry*; D008055:Lipids / Q000737:chemistry*; D008164:Luminescent Proteins / Q000378:metabolism; D008956:Models, Chemical; D010455:Peptides / Q000737:chemistry*; D010713:Phosphatidylcholines / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013347:Subcellular Fractions; D013696:Temperature
D003902:Detergents; D008051:Lipid Bilayers; D008055:Lipids; D008164:Luminescent Proteins; D010455:Peptides; D010713:Phosphatidylcholines; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D015060:1,2-Dipalmitoylphosphatidylcholine; D002784:Cholesterol; C017251:1,2-oleoylphosphatidylcholine