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234 Cell-Penetrating Peptides: Processes and Applications
10.6.1 STRUCTURE INDUCTION IN MICELLES
Detergent micelles, like mixed organic solvents, often induce helices in short peptides;
CPPs are no exception. For penetratin, the induction of α-helix has been shown
by NMR and CD in low (20 mM) and high (300 mM) SDS concentrations. 6,39 The
induction of secondary structure in peptides by these solvents is often explained in
terms of loss of hydrogen bonds from water.
There does not seem to be a close relationship between induction of α-helix by
these membrane-mimetic solvents and the membrane translocation property. In fact,
there are penetratin variants for which these solvents induce α-helical secondary
structures; yet they have lost the ability to translocate. Moreover, there are examples
of penetratin variants with proline insertions, which typically interrupt helical structures,
but these peptides are still able to translocate. Transportan in SDS micelles
gives a mixed structure: the C-terminal mastoparan part forms a well-developed
α-helix, whereas the galanin part has a less well-defined secondary structure. Both
parts are similar to structures of the corresponding parts of the galanin and mastoparan
peptides, respectively, in the same solvent. 24,40
10.6.2 POSITIONING IN MICELLES
In an attempt to carry NMR studies on peptides interacting with detergent micelles
one step further, recent studies have aimed at determining the positioning of different
parts of the peptide molecule relative to the surface and interior of the micelle. In
these studies paramagnetic probes are used, which affect the linewidths of the
resonances of nuclei in their neighborhood; these spin probes can be selected according
to where they reside in the micellar system. Spin-labeled fatty acids, such as
12-doxyl and 5-doxyl stearic acid, are known to act inside and at the surface of the
micelle, respectively. Paramagnetic ions, such as Mn 2+ or water-soluble spin labels,
affect parts of the peptide located outside the micelle. The selective effects on the peptide
resonances are investigated and information on peptide positioning is obtained. 41
Studies on transportan and penetratin in SDS micelles, using fatty acid spin
labels as internal and Mn 2+ ions as external paramagnetic probes, have been
reported. 39,42 The SDS was in large excess in these experiments, typically 300 mM
with 1 mM peptide. This corresponds to one peptide per five micelles, assuming 60
SDS per micelle. Results showed that the peptides are partly “hidden” inside the
micelle and partly exposed outside the micellar surface. The helical parts of transportan
and penetratin were found to be mostly buried by the micelle. For transportan,
the most C-terminal residues and the central segment connecting the galanin and
the mastoparan part were found to be exposed to the aqueous solvent. For penetratin,
the most N-terminal residues were found to be exposed to the aqueous solvent.
Therefore, a common denominator for transportan and penetratin in complex with
SDS micelles is that they both have a helix buried in the interior of the micelle and
an “anchor” at one or both ends of the helix.
The anchoring part is exposed at the outside of the micellar surface. This may
speculatively be regarded as a crude model of a CPP in contact with a membrane-like
environment with negative charge at the surface. The analysis of the shielding from the