Protein binding onto surfactant-based synthetic vesicles.
Synthetic vesicles were prepared by mixing anionic and cationic surfactants, aqueous sodium dodecylsulfate with didodecyltrimethylammonium or cetyltrimethylammonium bromide. The overall surfactant content and the (anionic/cationic) mole ratios allow one to obtain negatively charged vesicles. In the phase diagram, the vesicular region is located between a solution phase, a lamellar liquid crystalline dispersion, and a precipitate area. Characterization of the vesicles was performed by electrophoretic mobility, NMR, TEM, and DLS and we determined their uni-lamellar character, size, stability, and charge density. Negatively charged vesicular dispersions, made of sodium dodecylsulfate/didodecyltrimethylammonium bromide or sodium dodecylsulfate/cetyltrimethylammonium bromide, were mixed with lysozyme, to form lipoplexes. Depending on the protein/vesicle charge ratio, binding, surface saturation, and lipoplexes flocculation, or precipitation, occurs. The free protein in excess remains in solution, after binding saturation. The systems were investigated by thermodynamic (surface tension and solution calorimetry), DLS, CD, TEM, 1H NMR, transport properties, electrophoretic mobility, and dielectric relaxation. The latter two methods give information on the vesicle charge neutralization by adsorbed protein. Binding is concomitant to modifications in the double layer thickness of vesicles and in the surface charge density of the resulting lipoplexes. This is also confirmed by developing the electrophoretic mobility results in terms of a Langmuir-like adsorption isotherm. Charges in excess with respect to the amount required to neutralize the vesicle surface promote lipoplexes clustering and/or flocculation. Protein-vesicle interactions were observed by DLS, indicating changes in particle size (and in their distribution functions) upon addition of LYSO. According to CD, the bound protein retains its native conformation, at least in the SDS/CTAB vesicular system. In fact, changes in the alpha-helix and beta-sheet conformations are moderate, if any. Calorimetric methods indicate that the maximum heat effect for LYSO binding occurs at charge neutralization. They also indicate that enthalpic are by far the dominant contributions to the system stability. Accordingly, energy effects associated with charge neutralization and double-layer contributions are much higher than counterion exchange and dehydration terms.
Letizia
Caterina
C
Department of Chemistry, SOFT-INFM-CNR Research Centre, La Sapienza University, P. le A. Moro 5, I-00185 Rome, Italy.
Andreozzi
Patrizia
P
Scipioni
Anita
A
La Mesa
Camillo
C
Bonincontro
Adalberto
A
Spigone
Elisabetta
E
D000818:Animals; D000077286:Cetrimonium; D002593:Cetrimonium Compounds / Q000737:chemistry; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002645:Chickens; D004531:Eggs; D004563:Electrochemistry / Q000379:methods; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D046529:Microscopy, Electron, Transmission; D015233:Models, Statistical; D009113:Muramidase / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D013499:Surface Properties; D013501:Surface-Active Agents / Q000737:chemistry*
D002593:Cetrimonium Compounds; D013501:Surface-Active Agents; D012967:Sodium Dodecyl Sulfate; D009113:Muramidase; D000077286:Cetrimonium