Tilt and rotation angles of a transmembrane model peptide as studied by fluorescence spectroscopy.

In this study the membrane orientation of a tryptophan-flanked model peptide, WALP23, was determined by using peptides that were labeled at different positions along the sequence with the environmentally sensitive fluorescent label BADAN. The fluorescence properties, reflecting the local polarity, were used to determine the tilt and rotation angles of the peptide based on an ideal alpha-helix model. For WALP23 inserted in dioleoylphosphatidylcholine (DOPC), an estimated tilt angle of the helix with respect to the bilayer normal of 24 degrees +/- 5 degrees was obtained. When the peptides were inserted into bilayers with different acyl chain lengths or containing different concentrations of cholesterol, small changes in tilt angle were observed as response to hydrophobic mismatch, whereas the rotation angle appeared to be independent of lipid composition. In all cases, the tilt angles were significantly larger than those previously determined from (2)H NMR experiments, supporting recent suggestions that the relatively long timescale of (2)H NMR measurements may result in an underestimation of tilt angles due to partial motional averaging. It is concluded that although the fluorescence technique has a rather low resolution and limited accuracy, it can be used to resolve the discrepancies observed between previous (2)H NMR experiments and molecular-dynamics simulations.

Aggregation of transmembrane peptides studied by spin-label EPR.

The structure and function of membrane proteins is partly determined by the interaction of these proteins with the lipids of the membrane. Peptides forming single membrane-spanning alpha-helices, such as the WALP peptide (acetyl-GWWL(AL)(n)WWA-amide), are good models for such interactions. This interaction can be studied by investigating the aggregation of peptides. If the peptides remain isolated in the membrane, the peptide-lipid interaction dominates, if the peptides aggregate, the peptide-peptide interaction is stronger. The intrinsic dynamics and the disordered nature of the system require new approaches to determine eventual aggregation. We performed electron paramagnetic resonance (EPR) on spin-labeled WALP (SL-WALP) in the gel and the liquid-crystalline phases of two different phospholipids, the saturated DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine), and the unsaturated DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine). At low temperatures (120 K) where both lipids are in the gel phase, less extensive aggregation is observed for the peptide in DOPC as compared to DPPC. Together with previous data on aggregation of WALP peptides from atomic force microscopy and fluorescence spectroscopy at 294 K ( Sparr ; et al. Biochemistry 2005 , 44 , 2 -10 ), the results suggest that at 120 K WALP peptides form line aggregates in DOPC and cluster aggregates in DPPC. In the liquid-crystalline phase of both lipids, signatures of aggregation are absent, showing that in this phase the peptide can be accommodated by either lipid. It can be concluded that the lipid phase, in this case gel or liquid-crystalline, is a more important determinant for peptide aggregation than whether the lipid is saturated (DPPC) or unsaturated (DOPC). In view of the gel-phase-like behavior of some membrane phases in physiological systems the methodology should be relevant.

Synthesis and structural investigations of N-alkylated beta-peptidosulfonamide-peptide hybrids of the amyloidogenic amylin(20-29) sequence: Implications of supramolecular folding for the design of peptide-based bionanomaterials.

The incorporation of a single beta-aminoethane sulfonyl amide moiety in a highly amyloidogenic peptide sequence resulted in a complete loss of amyloid fibril formation. Instead, supramolecular folding morphologies were observed. Subsequent chemoselective N-alkylation of the sulfonamide resulted in amphiphilic peptide-based hydrogelators. It was found that variation of merely the alkyl chain induced a dramatic variation in aggregation motifs such as helical ribbons and tapes, ribbons progressing to closed tubes, twisted lamellar sheets and entangled/branched fibers.

Self-assembly of amylin(20-29) amide-bond derivatives into helical ribbons and peptide nanotubes rather than fibrils.

Uncontrolled aggregation of proteins or polypeptides can be detrimental for normal cellular processes in healthy organisms. Proteins or polypeptides that form these amyloid deposits differ in their primary sequence but share a common structural motif: the (anti)parallel beta sheet. A well-accepted approach for interfering with beta-sheet formation is the design of soluble beta-sheet peptides to disrupt the hydrogen-bonding network; this ultimately leads to the disassembly of the aggregates or fibrils. Here, we describe the synthesis, spectroscopic analysis, and aggregation behavior, imaged by electron microscopy, of several backbone-modified amylin(20-29) derivatives. It was found that these amylin derivatives were not able to form fibrils and to some extent were able to inhibit fibril growth of native amylin(20-29). However, two of the amylin peptides were able to form large supramolecular assemblies, like helical ribbons and peptide nanotubes, in which beta-sheet formation was clearly absent. This was quite unexpected since these peptides have been designed as soluble beta-sheet breakers for disrupting the characteristic hydrogen-bonding network of (anti)parallel beta sheets. The increased hydrophobicity and the presence of essential amino acid side chains in the newly designed amylin(20-29) derivatives were found to be the driving force for self-assembly into helical ribbons and peptide nanotubes. This example of controlled and desired peptide aggregation may be a strong impetus for research on bionanomaterials in which special shapes and assemblies are the focus of interest.