No abstract

Cooperative binding is a key feature of metabolic pathways, signaling, and transport processes. It provides tight regulation over a narrow concentration interval of a ligand, thus enabling switching to be triggered by small concentration variations. The data presented in this work reveal strong positive cooperativity of α-synuclein binding to phospholipid membranes. Fluorescence cross-correlation spectroscopy, confocal microscopy, and cryo -TEM results show that in excess of vesicles α-synuclein does not distribute randomly but binds only to a fraction of all available vesicles. Furthermore, α-synuclein binding to a supported lipid bilayer observed with total internal reflection fluorescence microscopy displays a much steeper dependence of bound protein on total protein concentration than expected for independent binding. The same phenomenon was observed in the case of α-synuclein binding to unilamellar vesicles of sizes in the nm and μm range as well as to flat supported lipid bilayers, ruling out that nonuniform binding of the protein is governed by differences in membrane curvature. Positive cooperativity of α-synuclein binding to lipid membranes means that the affinity of the protein to a membrane is higher where there is already protein bound compared to a bare membrane. The phenomenon described in this work may have implications for α-synuclein function in synaptic transmission and other membrane remodeling events.

In recent years, lipid bicontinuous cubic liquidcrystalline nanoparticles known as cubosomes have been under investigation because of their favorable properties as drug nanocarriers useful for anticancer treatments. Herein, we present organic/inorganic hybrid, theranostic cubosomes stabilized in water with a shell of alternate layers of chitosan, single strand DNA (model genetic material for potential gene therapy), and folic acid−chitosan conjugate (the outmost layer), coencapsulating up-converting Er 3+ and Yb 3+ codoped NaYF 4 nanoparticles and daunorubicin. The latter acts as a chemotherapeutic drug of photosensitizing activity, while up-converting nanoparticles serve as energy harvester and diagnostic agent. Cellular uptake and NIR-induced photodynamic therapy were evaluated in vitro against human skin melanoma (MeWo) and ovarian (SKOV-3) cancer cells. Results evidenced the preferential uptake of the theranostic cubosomes in SKOV-3 cells in comparison to uptake in MeWo cells, and this effect was enhanced by the folic acid functionalization of the cubosomes surface. Nanocarriers coloaded with the hybrid fluorophores exhibited a superior NIR-induced photodynamic activity, also confirmed by the improved mitochondrial activity and the most affecting f-actin fibers of cytoskeleton. Similar results, but with higher photocytotoxicity, were detected when folic acid-functionalized cubosomes were incubated with SKOV-3 cells. Taken on the whole, these results prove these hybrid cubosomes are good candidates for the photodynamic treatment of tumor lesions.