KEYWORDS: Fluorescence resonance energy transfer, Proteins, Luminescence, Quantum dots, Particles, Energy transfer, Molecules, Energy efficiency, Resonance energy transfer, Absorption
We present a single QD FRET study that allows us to probe the heterogeneity in QD-dye labeled protein conjugates. We first show that QDs are compatible with spFRET detection by demonstrating the equivalence of single particle and ensemble measurement modalities in terms of derived average FRET efficiencies and separation distances between a QD donor and dyes attached to specific sites on conjugated proteins. We then use spFRET data to demonstrate that the valence distribution of QD-protein conjugates follows Poisson statistics.
Vesicles are phospholipid bilayers that form a surface enclosing a volume of water or solution. They are of importance as model systems to study cells, as well as having practical applications such as containers for performing nanochemistry and facilitating drug delivery. Their properties have been studied for decades. Using a holographic laser tweezer array (LTA), which converts a single laser beam into many laser tweezer points, we stretch the vesicles in controlled ways from several points at once, measuring each force applied. Here, we present data on shape deformations of simple, spherical vesicles and on membrane fracture.
We have proposed using single molecule fluorescence resonant energy transfer (SM-FRET) to investigate the induction of secondary structure in model, surface-active peptides upon binding at an interface. The ability for SM-FRET to distinguish structural heterogeneity will offer a distinct advantage over traditional biophysical methods in these types of studies. Ensemble methods mask heterogeneity and only provide an average measure of secondary structural features. Because secondary structure contributes greatly to the energetics of dehydrating the amide backbone, detailed information of conformational distributions is crucial to the understanding of the thermodynamic cycle involved. Here we present results from our first efforts at using SM-FRET to study an amphipathic α-helix forming peptide immobilized at the solid-liquid interface between an aqueous solution and an octadecylsilane modified glass surface. This system serves as a model for future studies of peptide partitioning to lipid bilayers and other relevant interfaces.
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