Biophysical Society Thematic Meeting

Liposomes, Exosomes, and Virosomes: From Modeling Complex

Membrane Processes to Medical Diagnostics and Drug Delivery

Monday Speaker Abstracts

Use of Model Membranes-Liposomes and Micelles of Variable Lipid Composition to

Elucidate the Molecular Mechanism of Action of Pore-forming Proteins and Peptides

Gustavo P. Carretero

, Eduardo M. Cilli

, Carlos Alvarez

Shirley Schreier

University of Sao Paulo, Sao Paulo, Sao Paulo, Brazil,

State University of Sao Paulo,

Araraquara, Sao Paulo, Brazil,

University of La Habana, La Habana, Cuba.

Sticholysins I and II, cytolysins purified from the sea anemone Stichodactyla helianthus, lyse

biological and model membranes. The proposed mechanism of action consists of the formation

of a toroidal pore involving the N-terminal domain. The interaction between peptides from the

toxins’ N-termini (StI1-31 and StI12-31 SELAGTIIDGASLTFEVLDKVLGELGKVSRK, and

StII1-30 and StII11-30 ALAGTIIAGASLTFQVLDKVLEELGKVSRK) and model membranes

– liposomes and micelles – was studied in order to contribute to the elucidation of the toxins

mechanism of action at the molecular level. Peptides were used based on the hypothesis that

protein fragments can mimic the structure and activity of the whole protein. An analogue

containing the paramagnetic amino acid TOAC (N-TOAC-StII11-30) was also studied.

Conformational studies made use of circular dichroism (CD), electron paramagnetic resonance

(EPR), and fluorescence. Studies of structure prediction and molecular modeling were also

performed. The peptides acquired α-helical conformation upon interaction with model lipid

membranes, in agreement with the conformation found for these segments in the whole proteins.

Studies with membranes of variable lipid composition, demonstrated that both electrostatic and

hydrophobic interactions contribute to peptide binding. Fluorescence quenching of labeled lipids

by paramagnetic TOAC and EPR spectra allowed us to locate the TOAC residue at the

membrane-water interface, corroborating the proposed model of the toroidal pore. CD and EPR

studies also allowed calculation of peptide-membrane binding constants. The peptides also

mimicked the toxins function, as shown by assays of carboxyfluorescein leakage and hemolytic

activity. Short peptides containing parts of StII1-30’s sequence were synthesized with the aim of

testing their antimicrobial activity. The peptides displayed low antimicrobial activity, as well as

lack of hemolytic activity and toxicity against human cells.