90 / 92
- 89 -
Polymers and Self Assembly: From Biology to Nanomaterials Poster Session II
43-POS
Board 43
Synthesis, Characterization and Applications of Size Restricted Polycationic Hen Lysozyme
Nanoparticles
Tulsi Swain
.Rajaram Swaminathan,
Indian Inst. Tech. Guwahati, Guwahati, Assam, India.
We report a novel strategy for synthesis of Hen Egg-White Lysozyme (HEWL) nanoparticles by
alkaline pH-induced aggregation at room temperature with subsequent stabilization by
intermolecular disulfide bonds [1]. Size heterogeneity of the nanoparticles renders their
characterization a formidable challenge. To address this we have used size exclusion and ion
exchange chromatography to separate differently sized nanoparticles. Further biophysical
methods like steady state fluorescence anisotropy, DLS, TEM, AFM and SAXS were employed
to characterize the size-fractionated nanoparticles in detail. Accessible hydrophobic regions in
the HEWL nanoparticles were assessed by ANS fluorescence. Pyrene fluorescence sensitivity to
solvent polarity has been used to estimate changes in probe microenvironment as a result of
conformational change during aggregation of HEWL. Significant drop in steady state anisotropy
of fluorescein conjugated fractions clearly shows that aggregates labeled with multiple
fluorophores exhibit homo-FRET. Self-quenching of fluorescein limits the brightness available
per labeled protein. But in our case the phenomenon of self-quenching is turned into an
advantage to study aggregation. The fluorescein conjugated nanoparticles can be used for
targeted molecular imaging by attaching appropriate ligands to them. Polycationic HEWL
nanoparticles have numerous positive charges on their surface which can facilitate association of
polyplexes to various negatively charged cargoes.These nanoparticles were able to condense
DNA efficiently as revealed from gel retardation assay. The promising HEWL nanoparticles can
be utilized as imaging nanovectors, DNA delivery vehicle and F19 contrast agents. These
nanoparticles also offer the advantage of being engineered for use as drug delivery vehicle in
future.
[1] Ravi VK, Swain T, et. al., (2014) PLoS ONE 9(1): e87256.
doi:10.1371/journal.pone.0087256