Formation of Lysozyme Oligomers at Model Cell Membranes Monitored with Sum Frequency Generation Spectroscopy

Abstract

A growing number of studies suggest that the formation of toxic oligomers, precursors of amyloid fibrils, is initiated at the cell membrane and not in the cytosolic compartments of the cell. Studies of membrane-induced protein oligomerization are challenging due to the difficulties of probing small numbers of proteins present at membrane surfaces. Here, we employ surface-sensitive vibrational sum frequency generation (VSFG) to investigate the secondary structure of lysozyme at the surface of lipid monolayers. We investigate lysozyme aggregation at negatively charged 1,2-dipalmitoyl-sn-glycero-3-(phospho-rac-1-glycerol) (DPPG) lipid monolayers under different pH conditions. The changes in the molecular vibrations of lipids, proteins, and water as a function of pH and surface pressure allow us to simultaneously monitor details of the conformation state of lysozyme, the organization of lipids, and the state of lipid-bound water. At pH = 6 lysozyme induces significant disordering of the lipid layer, and it exists in two states: a monomeric state with a predominantly α-helix content and an oligomeric (za-mer) state. At pH ≤ 3, all membrane-bound lysozyme self-associates into oligomers characterized by an antiparallel β-sheet structure. This is different from the situation in bulk solution, for which circular dichroism (CD) shows that the protein maintains an α-helix conformation, under both neutral and acidic pH conditions. The transition from monomers to oligomers is also associated with a decreased hydration of the lipid monolayer resulting in an increase of the lipid acyl chains ordering. The results indicate that oligomerization requires cooperative action between lysozyme incorporated into the lipid membrane and peripherally adsorbed lysozyme and is associated with the membrane dehydration and lipid reorganization. Membrane-bound oligomers with antiparallel β-sheet structure are found to destabilize lipid membranes.

SAP(E) – A cell-penetrating polyproline helix at lipid interfaces

“…Model membranes simplify investigations of specific peptide-membrane interactions by providing a defined environment. Lipid monolayers are a well-established model membrane system [37] that has been widely used to get information about peptide-membrane interactions [38][39][40][41][42]. We employed saturated lipids, for experimental reasons, with different headgroup charges, i.e.…”

Bovine and human insulin adsorption at lipid monolayers: a comparison

“…Based on works by Brange et al it is unlikely that human insulin will aggregate more readily in solution than bovine insulin, it is likely that such an oligomerization would be initiated by the presence of the lipid interface [24]. Such surface induced oligomerization has been observed before [14,18].…”

“…Recent works have shown, that fibril formation can be initiated by the presence of lipid surfaces [13][14][15]. The interaction of insulin with biologically relevant lipid membranes surfaces has not been investigated yet.…”

Sticky water surfaces: Helix–coil transitions suppressed in a cell-penetrating peptide at the air-water interface

“…34 SFG has been successfully used to probe pro- tein and peptide secondary structures in the past. In those SFG studies α-helix, [35][36][37] β-sheet, 35, 37 3 10 -helix, 38 and protein aggregates [39][40][41] have been identified. Figure 3(a) shows a scheme of the experimental setup.…”