10.1002/chem.201805880
Chemistry - A European Journal
COMMUNICATION
interface when compared to P2 and P3. The origins of an
additional distance in P2, confirmed by a validation analysis (Fig.
S7b), is still unclear but do not interfere with the overall
interpretation of the results.
Acknowledgements
We thank B. Angerstein for help with preparation of lipid samples.
We acknowledge financial support from the Deutsche
Forschungsgemeinschaft (DFG) collaborative research program
SFB 803 (project A02), from the Georg-August University of
Göttingen and the Max Planck Society.
The determined peak distances enabled to examine the helical
structure of the investigated peptide in more details. Simplified
models of four doubly-labeled peptides based on 314-helices were
constructed and corresponding inter-spin distances extracted
(Table S5‒8). Coordinates of the backbone atoms were
generated from torsional angles known in literature (Table
S4).[22,23] Since data for torsional angles of 314-helices differ in
literature, idealized model peptides were generated according to
two backbone angle sets named as 3.2514 and 3.014 (models are
illustrated in Figure 2 A and Figure S8‒9). For the individual
peptides, the inter-spin distances gained from the models were
compared with the experimental ones by plotting the distances
against the number of amino acids sequentially separating the two
labels (Δaa) (Figure 2 B). The obtained curves indicate a better
agreement of the experimental data with the structure created
from the angle set 3.2514 rather than 3.014. In addition, the peak
distances extracted from model 3.2514 and the ones measured
with the PELDOR experiment are in close agreement, in solution
as well as in the lipid environment. Small deviations between
predicted and experimental data most likely result from the
structural simplification and the disregard of peptide-environment
interactions in the model. This implies that a helical turn of the β3-
peptides is defined by approximately 3.25 amino acid residues,[23]
which is supported by a NMR study of a β3-eicosapeptide
consisting of homologated proteinogenic amino acids in MeOH by
Seebach and co-workers.[24] According to the results from
PELDOR experiments in MLVs, the -peptide helix conformation
is preserved in the lipid environment. From the PELDOR
experiments it can be concluded that the labeled β3-peptides P2‒
P5, and thus the reference peptide P1, fold into a 3.2514-helix that
is conformationally stable in solution and in the lipid bilayer.
In conclusion, we have demonstrated that the combination of a
new sophisticated spin label, such as the novel semi-rigid β3-
hTOPP label, with EPR and PELDOR spectroscopy provides a
powerful and straightforward tool for the structural investigation of
transmembrane β-peptides. Due to the semi-rigid design of the
β3-hTOPP label, site-specific distance measurements allowed for
investigation of the peptide’s backbone conformation with
resolution at the molecular length scale even in a lipid
environment. Comparison of the experimentally determined peak
distances with structural models of -peptides revealed that the
oligomers fold into a 3.2514-helix rather than an idealized 3.014-
helical conformation. Since this method was also shown to be
transferable to hydrophobic transmembrane conditions, it serves
as a promising tool for the structural analysis of transmembrane
-peptides and membrane protein domains in case they are
modified with -peptide secondary structure mimics.
Keywords: -Peptides • EPR spectroscopy • membrane
proteins • nitroxide radicals • PELDOR • spin labels
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Experimental Section
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