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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
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