Optical Reporting by Conjugated Polymers via Conformational Changes - Advanced Fluorescence Reporters in Chemistry and Biology - page 408

a b

7000

Buffer
Native insulin
Armyloid insulin

6000

5000

4000

3000

2000

1000

0
480 530 580 630 680

Wavelength (nm)

Fig. 17 (a) Chemical structure of polythiophene poly((3,3 00 -di[(S)-5-amino-5-carbonyl-3-oxapen-
tyl]-[2,2 0 ;5 0 2 00 ])-5-,5 00 -terthiophenylene hydrochloride), PONT. (b) Emission spectra of 6.5 m M

PONT HCl (on a chain basis) in 25 mM HCl ( black spectrum ), 25 mM HCl with 5.0 m M of native

bovine insulin ( blue spectrum ), 25 mM HCl with 5.0 M fibrillar bovine insulin ( red spectrum ).

The emission spectra were recorded with excitation at 400 nm [ 31 ]

m

light at these wavelengths, Ration 550/580 nm, the kinetics of the fibrillation event
could be followed (Fig. 16c ). The ability to detect amyloid fibrils was demonstrated
with both bovine insulin and chicken lysozyme to verify PTAAs usefulness as a
conformation-sensitive probe. Hence, the PTAA emission profile was specific
for the repetitive

b sheet structure independent of the primary sequence of the

protein used.
Later work, by Herland et al. synthesized a well-defined zwitterionic polythio-
phene, PONT, (Fig. 17a )[ 31 ]. On the basis of the previous work with serine substi-
tuted polythiophenes [ 9 ], Herland et al. synthesized regio-regular, well-defined
molecules by chemical oxidation, which resulted in zwitter-ionic oligomeric thio-
phene derivatives from about 9 to 15 monomer units in length. This reduction in the
distribution of chain lengths and polydispersity in the polymers helps to increase
specificity of the probes and clarity in the interpretation of fluorescence data. Their
application of choice was for the detection of amyloid fibrils of bovine insulin and
chicken lysozyme. As shown in Fig. 17b , PONT showed a specific emission profile
bound to amyloid fibrils, whereas the spectra for the polymer free in solution or
mixed with native insulin were similar. The increased intensity of the emission and
the blue-shift of the emission maximum upon binding to the amyloid fibrils are
mainly due to separation of the polymer chains, whereas the decreased intensity and
red-shift of the emission seen for PONT in acidic solvents or mix with native insulin
correspond to planarization of the backbone, which causes aggregation or stacking
of the polymers and interchain de-excitation between polymer chains. Hence, the
authors were able to show that the regio-regular polymers could detect and identify
amyloid fibrils specifically and reliably under the acidic conditions often required
for in vitro fibrillization protocols. A recent study, comparing the amyloid selectiv-
ity of polythiophenes synthesized from monomeric or trimeric building blocks, has