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Luminescent Conjugated Polymers for Staining and Characterization of Amyloid Deposits 339
changes considerably, resulting in a rearrangement from a closed to
an open conformation of the protein motif. Structural studies also
suggest that calcium activation of the protein is accompanied by a
global conformational change, whereby the compact calcium-free
form of CaM is converted to a more extended dumbbell-shaped mol-
ecule upon binding of calcium. 90, 91 The extended form of the protein
consists of two lobes separated by a central α helix, and this central
helix is flexible and allows considerable movements of the two lobes
with respect to each other.
Upon formation of a complex between POWT and CaM, the emis-
sion maximum of POWT is red-shifted and the intensity of the emitted
light is decreased (Fig. 9.5b) (relative to POWT alone in the same buffer),
indicating that the POWT backbone becomes more planar and that
2+
aggregation of the POWT chains occurs. An addition of 10 mM Ca to
this complex will blue shift the emission maximum (594 nm), and the
shoulder around 540 nm is increasing (Fig. 9.5b), suggesting that the
polymer backbone becomes more nonplanar and that a separation of
the polymer chains occurs. The ratio of the intensity of the emitted light
at 540/670 nm is increased, showing that the conformational change of
2+
the CaM molecule upon exposure to Ca is governing the geometry of
the POWT chains. The increased emission at 540 nm, associated with
separation of the polymer chains, is probably a result of the conforma-
tional changes of CaM that occur upon binding of calcium. A schematic
presentation of the different conformational alterations of the CaM
molecule upon exposure to calcium and the suggested POWT chain
geometries interpreted from the spectral changes seen for the different
POWT/CaM solutions is shown in Fig. 9.5c. 86
As discussed above, it is rather evident that the conformation-
sensitive optical properties of LCPs can be used as an optical finger-
print for distinct protein conformations. Hence, LCPs can be applied
as a novel tool within the research field of protein folding and protein
aggregation diseases. The underlying mechanism of protein aggrega-
tion, e.g., the formation of amyloid fibrils, and the diseases believed
to be associated with this event are discussed in greater detail next.
9.3 Amyloid Fibrils and Protein Aggregation Diseases
9.3.1 Formation of Amyloid Fibrils
Proteins frequently alter their conformation due to different external
stimuli, and many diseases are associated with misfolded proteins. 92, 93
Especially under conditions that destabilize the native state, proteins
can self-assemble into aggregated β-sheet rich fibrillar assemblies,
known as amyloid fibrils, which are around 10 nm wide and unusually
stable biological materials (Fig. 9.6). However, the process where by a
native protein is converted to amyloid fibrils is quite complex, and