Fluorescent Environment-Sensitive Dyes as Reporters of Biomolecular Interactions - Progress in Molecular Biology and Translational Science - page 53

of this process and detection of the aggregated structures much earlier than
other techniques.
4.4. Peptide - membrane interactions
Monitoring interactions of proteins with biological membranes is of partic-
ular importance for studying membrane proteins and peptide-based toxins
and for understanding the peptide transport through cell membranes. These
interactions should lead to remarkable changes in the polarity of the peptide
environment, as the rather polar peptide interface is substituted with the
highly apolar lipid membrane environment. Therefore, the 3HC-A dye is
more appropriate for these studies. Its carboxylic acid derivative was attached
to the N-terminus of melittin and poly- L -lysine peptides, which interact
with lipid membranes in a very different manner. Binding of these peptides
to lipid vesicles induced a strong fluorescence increase, which enabled the
quantification of the peptide-membrane interactions. 66 Moreover, the dual
emission of the label in these peptides correlated well with the depth of its
insertion, measured by the parallax quenching method ( Fig. 2.10 ). Thus, in
melittin, which shows deep insertion of its N-terminus, the label presented a
dual emission corresponding to a low-polar environment, while the envi-
ronment of the poly- L -lysine N-terminus was rather polar, in line with its
binding at the surface of the lipid head groups. Moreover, imaging of labeled
peptides bound to giant vesicles gave some clues on the orientation of the
label within the membrane, which could help estimating the peptide
orientation.
This label was further successfully applied for monitoring the interaction
of a -synuclein with model membranes, as this interaction was hypothesized
to play a role in the pathological misfolding and aggregation of this protein
during Parkinson's disease. Systematic studies of a -synuclein labeled with
3HC-A fluorophore revealed the influence of charge, phase, curvature,
defects, and lipid unsaturation on binding of a -synuclein to model
membranes 67 and its further conformational changes. 68
5. CONCLUSIONS
Solvatochromic dyes, because of their ability to undergo excited-state
reactions (charge and proton transfer), can change their emission color and
intensity in response to variation of solvent polarity. Though a number of