Poly-l-lysines and poly-l-arginines induce leakage of negatively charged phospholipid vesicles and translocate through the lipid bilayer upon electrostatic binding to the membrane.
Poly-l-lysines (PLL) and poly-l-arginines (PLA) of different polymer chain lengths interact strongly with negatively charged phospholipid vesicles mainly due to their different electrical charges. 1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG), 1,2-dipalmitoyl-sn-glycero-3-phosphoglycerol (DPPG) and their mixtures (1/1 mol/mol) with the respective phosphatidylcholines of equivalent chain length were chosen as model membrane systems that form at room temperature either the fluid L(alpha) or the gel phase L(beta) lipid bilayer membranes, respectively. Leakage experiments revealed that the fluid POPG membranes are more perturbed compared to the gel phase DPPG membranes upon peptide binding. Furthermore, it was found that pure PG membranes are more prone to release the vesicle contents as a result of pore formation than the lipid mixtures POPG/POPC and DPPG/DPPC. For the longer polymers (>or=44 amino acids) maximal dye-release was observed when the molar ratio of the concentrations of amino acid residues to charged lipid molecules reached a value of R(P)=0.5, i.e. when the outer membrane layer was theoretically entirely covered by the polymer. At ratios lower or higher than 0.5 leakage dropped significantly. Furthermore, PLL and PLA insertions and/or translocations through lipid membranes were analyzed by using FITC-labeled polymers by monitoring their fluorescence intensity upon membrane binding. Short PLL molecules and PLA molecules of all lengths seemed to translocate through both fluid and gel phase lipid bilayers. Comparison of the PLL and PLA fluorescence assay results showed that PLA interacts stronger with phospholipid membranes compared to PLL. Isothermal titration calorimetry (ITC) measurements were performed to give further insight into these mechanisms and to support the findings obtained by fluorescence assays. Cryo-transmission electron microscopy (cryo-TEM) was used to visualize changes in the vesicles' morphology after addition of the polypeptides.
Reuter
Marcel
M
Institute of Chemistry, Martin-Luther-University Halle-Wittenberg, 06108 Halle, Germany.
Schwieger
Christian
C
Meister
Annette
A
Karlsson
Göran
G
Blume
Alfred
A
D020285:Cryoelectron Microscopy; D005456:Fluorescent Dyes / Q000737:chemistry; D008051:Lipid Bilayers / Q000737:chemistry*; D008081:Liposomes / Q000737:chemistry; D008567:Membranes, Artificial; D010455:Peptides / Q000737:chemistry*; D010715:Phosphatidylglycerols / Q000737:chemistry; D011107:Polylysine / Q000737:chemistry*; D055672:Static Electricity
D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D008081:Liposomes; D008567:Membranes, Artificial; D010455:Peptides; D010715:Phosphatidylglycerols; D011107:Polylysine; C015462:polyarginine; C060037:1-palmitoyl-2-oleoylglycero-3-phosphoglycerol; C030345:1,2-dipalmitoylphosphatidylglycerol