INSERTION IN LIPID MONOLAYERS OF ST II, A PORE-FORMING TOXIN, IS MODULATED BY THE PRESENCE OF CHOLESTEROL AND SPHINGOMYELIN

MARTINEZ, D.(1), ALVAREZ, C. (1), LANIO, M.E. (1), PAZOS, F. (1), TEJUCA, M. (1), GUTIERREZ-AGUIRRE, I.(2), GONZALEZ-MANAS, J.M. (2).

(1)University of Havana, Cuba, (2)The University of Basque Country, Spain.

Sticholysin II (St II) is a polypeptide toxin produced by the sea anemone Stichodactyla helianthus characterized by forming pores in membranes and its activity is inhibited by sphingomyelin. The interaction of this toxin with lipid monolayers was determined by measuring changes in surface pressure as a function of the lipid composition, particularly the influence of phosphatidyl choline (PC), sphingomyelin(SM), cholesterol (Cho) and their mixtures were thoroughly studied.  Association of St II to single lipids was analyzed in monolayers as well as in an ELISA with lipid coating, revealing in both systems a remarkable preference for SM, two fold higher than that obtained for PC. Using monolayers comprising binary lipid mixtures, the highest critical pressure values (p_{c) were reached for SM:Cho mixtures close to equimolarity while the lowest values were attained for mixtures that did not contain SM. Ternary mixtures containing 50% SM and more than 30% cholesterol resulted optimal for the toxin, considering their high} p_{c values and the slope of the curves} Dp/p_{0. Insertion kinetics in monolayers, at the same starting pressure, as a function of lipid composition evidenced differences in the extreme composition cases (PC, PC:Cho and andall the ternary mixtures containing 50 % SM and 30 % Cho) both in terms of the process extension as well as rate. This phenomenon is coincident with that observed in permeabilization assays of LUV loaded with fluorophores. This behavior could be explained by the formation of lipid domains facilitating the toxin insertion even though a better accommodation of the protein can not be ruled out.}

CORRESPONDENCE TO:

Carlos Alvarez, Centro de Estudio de proteínas, Facultad de Biología, Universidad de la Habana, Calle 25, entre J e I, Vedado, Ciudad Habana, Cuba, Email: calvarez@infomed.sld.cu