amyloid-beta-peptides and 1-palmitoyl-2-oleoylglycero-3-phosphoglycerol

Short range plasmonic fields around a nanoparticle can modulate fluorescence or Raman processes. In lipid encased nanoparticles, this can potentially measure the relative depths of different parts of a membrane protein from the surface. We employ this technique to discover that membrane inserted amyloid-β oligomers have a preferred molecular orientation.

Topics: Amyloid beta-Peptides; Cholesterol; Fluoresceins; Fluorescence; Fluorescent Dyes; Lipid Bilayers; Metal Nanoparticles; Peptide Fragments; Phosphatidylcholines; Phosphatidylglycerols; Silver; Spectrometry, Fluorescence; Spectrum Analysis, Raman

Small hydrophobic oligomers of aggregation-prone proteins are thought to be generically toxic. Here we examine this view by perturbing an early folding contact between Phe19 and Leu34 formed during the aggregation of Alzheimer's amyloid-β (Aβ40) peptide. We find that even conservative single mutations altering this interaction can abolish Aβ40 toxicity. Significantly, the mutants are not distinguishable either by the oligomers size or by the end-state fibrillar structure from the wild type Aβ40. We trace the change in their toxicity to a drastic lowering of membrane affinity. Therefore, nonlocal folding contacts play a key role in steering the oligomeric intermediates through specific conformations with very different properties and toxicity levels. Our results suggest that engineering the folding energy landscape may provide an alternative route to Alzheimer therapeutics.

Topics: Amyloid beta-Peptides; Animals; Cell Survival; Cells, Cultured; Cerebral Cortex; Membranes, Artificial; Mutation; Neurons; Peptide Fragments; Phosphatidylcholines; Phosphatidylglycerols; Protein Folding; Rats, Wistar; Unilamellar Liposomes

Beta-amyloid peptide (Abeta) is a primary protein component of senile plaques in Alzheimer's disease (AD) and plays an important, but not fully understood role in neurotoxicity. Model peptides with the demonstrated ability to mimic the structural and toxicity behavior of Abeta could provide a means to evaluate the contributions to toxicity that are common to self-associating peptides from many disease states. In this work, we have studied the peptide-membrane interactions of a model beta-sheet peptide, P(11-2) (CH(3)CO-Gln-Gln-Arg-Phe-Gln-Trp-Gln-Phe-Glu-Gln-Gln-NH(2)), by fluorescence, infrared spectroscopy, and hydrogen-deuterium exchange. Like Abeta(1-40), the peptide is toxic, and conditions which produce intermediate oligomers show higher toxicity against cells than either monomeric forms or higher aggregates of the peptide. Further, P(11-2) also binds to both zwitterionic (POPC) and negatively charged (POPC:POPG) liposomes, acquires a partial beta-sheet conformation in presence of lipid, and is protected against deuterium exchange in the presence of lipids. The results show that a simple rationally designed model beta-sheet peptide recapitulates many important features of Abeta peptide structure and function, reinforcing the idea that toxicity arises, at least in part, from a common mode of action on membranes that is independent of specific aspects of the amino acid sequence. Further studies of such well-behaved model peptide systems will facilitate the investigation of the general principles that govern the molecular interactions of aggregation-prone disease-associated peptides with cell and/or membrane surfaces.

Topics: Amyloid beta-Peptides; Cell Line, Tumor; Cell Survival; Deuterium Exchange Measurement; Humans; Liposomes; Mass Spectrometry; Peptide Fragments; Peptides; Phosphatidylglycerols; Spectroscopy, Fourier Transform Infrared

The beta-amyloid peptide beta AP(1-40), a 40-amino acid residues peptide, is one of the major components of Alzheimer's amyloid deposits. beta AP(1-40) exhibits only a limited solubility in aqueous solution and undergoes a concentration-dependent, cooperative random coil reversible beta-structure transition for Cpep > 10 microM [Terzi, E., Hölzemann, G., and Seelig, J. (1995) J. Mol. Biol. 252, 633-642]. In the presence of acidic lipid, the equilibrium is shifted further toward beta-structured aggregates. We have now characterized the lipid-peptide interaction using circular dichroism (CD) spectroscopy, lipid monolayers, and deuterium and phosphorus-31 solid-state nuclear magnetic resonance (NMR). CD spectroscopy revealed a distinct interaction between beta AP(1-40) and negatively charged unilamellar vesicles. In addition to the random coil reversible beta-structured aggregate equilibrium at low lipid-to-peptide (L/P) ratios, a beta-structure -->alpha-helix transition was observed at L/P > 55. beta AP(1-40) was found to insert into acidic monolayers provided the lateral pressure was low (20 mN/m). The extent of incorporation increased distinctly with the content of acidic lipid in the monolayer. However, at a lipid packing density equivalent to that of a bilayer (lateral pressure > or = 32 mN/m), no insertion of beta AP(1-40) was observed. The lipid molecular structure in the presence of beta AP(1-40) was studied with NMR. Phosphatidylcholine (PC) was selectively deuterated at the choline headgroup and at the cis-double bond of the oleic acyl chain and mixed with phosphatidylglycerol (PG). Phosphorus-31 NMR showed that the lipid phase retained the bilayer structure at all lipid-to-protein ratios. Deuterium NMR revealed no change in the headgroup conformation of the choline moiety or in the flexibility and ordering of the hydrocarbon chains upon the addition of beta AP-(1-40). It can be concluded that beta AP(1-40) binds electrostatically to the outer envelope of the polar headgroup region without penetrating between the polar groups. The data suggest a new mechanism of helix formation induced by the proper alignment of five positive charges of beta AP(1-40) on the negatively charged membrane template.

Topics: Amino Acid Sequence; Amyloid beta-Peptides; Circular Dichroism; Deuterium; Membranes, Artificial; Molecular Sequence Data; Nuclear Magnetic Resonance, Biomolecular; Peptide Fragments; Phosphatidylcholines; Phosphatidylglycerols; Phosphorus Isotopes; Protein Binding; Protein Structure, Secondary