Cat-anionic vesicles are aggregates formed by mixing two oppositely charged surfactants, in non-stoichiometric ratios [1,2]. Their structure is similar to cellular membranes and, for this reason, they could be employed as vectors for bio-active molecules in drug delivery technologies and transfection, particularly in non-viral gene therapy. For this reason, it is fundamental to study their interaction with biomacromolecules. Vesicular entities were obtained by mixing didodecyldimethylammonium bromide and sodium dodecylsulfate in non-stoichiometric [DDAB]/[SDS] ratios (3.8) and the total surfactant concentration is 4 mmol kg-1. Vesicles bear a positive surface charge, due to the cationic species in excess. In our study, the binding of bovine serum albumin (BSA) was studied. At its spontaneous pH, BSA has a negative effective charge. In these conditions, vesicles adsorb significant amount of protein by electrostatic interactions, presumably. Moreover, we modulated the bovine serum albumin net charge by pH and dealt with its binding onto the above vesicles. Binding is controlled by the net charge of vesicles and albumin: it is substantial when albumin has negative charges in excess and is negligible, if any, below its isoelectric point. For pH > 6.0, the binding efficiency increases in proportion to protein charge. Surface coverage changes in proportion to pH, when the number of charges neutralized upon binding remains the same. The size of protein-vesicle lipo-plexes was inferred by Dynamic Light Scattering and their charge by ζ-potential. The structure of albumin was evaluated by Circular Dichroism spectroscopy and estimates of α-helix, β-strand and random coil content were achieved. Increasing of β-strand and random coil content subsequent to binding suggests a strong interaction between vesicles and albumin. Attempts to determine the binding efficiency were made by elaborating surface charge density values by ζ-potential ones. The results were interpreted in terms of a Gibbs adsorption isotherm. Accordingly, it is possible calculating the binding energy in different pH conditions. The results obtained so far indicate the existence of strong electrostatic interactions between protein and vesicles with fast formation of stable complexes, and highlight the pontentialites of these cat-anionic vesicles in nanomedicine. 1. Kaler E.W., Murthy A.K., Rodriguez B.E., Zasadzinski J.A., Science 1989, 245, 1371. 2. Yatcilla M.T., Herrington K.L., Brasher L.L., Kaler E.W., Chiruvolu S., Zasadzinski J.A., J. Phys. Chem. 1996, 100, 5874. 3. Ulrich, A.S., Biosci. Rep. 2002, 22, 129.
Study of protein binding onto cat-anionic vesicles for nanomedicine applications / Pucci, Carlotta; Scipioni, Anita; LA MESA, Camillo. - STAMPA. - (2012). (Intervento presentato al convegno 2nd international conference on Nanomedicine - from molecules to diagnosis and therapy tenutosi a Roma nel 1-3 Ottobre 2012).
Study of protein binding onto cat-anionic vesicles for nanomedicine applications
PUCCI, CARLOTTA;SCIPIONI, Anita;LA MESA, Camillo
2012
Abstract
Cat-anionic vesicles are aggregates formed by mixing two oppositely charged surfactants, in non-stoichiometric ratios [1,2]. Their structure is similar to cellular membranes and, for this reason, they could be employed as vectors for bio-active molecules in drug delivery technologies and transfection, particularly in non-viral gene therapy. For this reason, it is fundamental to study their interaction with biomacromolecules. Vesicular entities were obtained by mixing didodecyldimethylammonium bromide and sodium dodecylsulfate in non-stoichiometric [DDAB]/[SDS] ratios (3.8) and the total surfactant concentration is 4 mmol kg-1. Vesicles bear a positive surface charge, due to the cationic species in excess. In our study, the binding of bovine serum albumin (BSA) was studied. At its spontaneous pH, BSA has a negative effective charge. In these conditions, vesicles adsorb significant amount of protein by electrostatic interactions, presumably. Moreover, we modulated the bovine serum albumin net charge by pH and dealt with its binding onto the above vesicles. Binding is controlled by the net charge of vesicles and albumin: it is substantial when albumin has negative charges in excess and is negligible, if any, below its isoelectric point. For pH > 6.0, the binding efficiency increases in proportion to protein charge. Surface coverage changes in proportion to pH, when the number of charges neutralized upon binding remains the same. The size of protein-vesicle lipo-plexes was inferred by Dynamic Light Scattering and their charge by ζ-potential. The structure of albumin was evaluated by Circular Dichroism spectroscopy and estimates of α-helix, β-strand and random coil content were achieved. Increasing of β-strand and random coil content subsequent to binding suggests a strong interaction between vesicles and albumin. Attempts to determine the binding efficiency were made by elaborating surface charge density values by ζ-potential ones. The results were interpreted in terms of a Gibbs adsorption isotherm. Accordingly, it is possible calculating the binding energy in different pH conditions. The results obtained so far indicate the existence of strong electrostatic interactions between protein and vesicles with fast formation of stable complexes, and highlight the pontentialites of these cat-anionic vesicles in nanomedicine. 1. Kaler E.W., Murthy A.K., Rodriguez B.E., Zasadzinski J.A., Science 1989, 245, 1371. 2. Yatcilla M.T., Herrington K.L., Brasher L.L., Kaler E.W., Chiruvolu S., Zasadzinski J.A., J. Phys. Chem. 1996, 100, 5874. 3. Ulrich, A.S., Biosci. Rep. 2002, 22, 129.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
