Elizabeth Plum, Biochemistry and Molecular Biology
"Aggregation of Potential Drug Delivery Vehicle"
Faculty Mentor: Dr. Richard Karpel
Expected Graduation Date: May 2010
Rattlesnakes are well known for their poisonous bites but surprisingly, the venom of the South American rattlesnake Crotalus durissus terricus, possesses a valuable protein called crotamine. This protein is a 42- residue polypeptide with unusual properties not known to other toxins. It is a cell penetrating protein (CPP) and once inside cells, it localizes on chromosomes. Crotamine is not only able to penetrate, but is also a transporter, able to carry small molecules or even entire genes into cells. For example, it has been previously shown that crotamine facilitated the transfection of plasmid DNA into mice bone marrow. Crotamine specifically enters only actively proliferating (AP) cells through an interaction with heparin-glycosaminoglycans (GAG). It forms ternary complexes with these GAGs and DNA. Crotamine is able to bind to negatively charged DNA because it is highly basic, containing 9 lysine and 2 arginine residues. Crotamine-DNA complexes then enter the cell by endocytosis and localize on the nucleus. Past experiments I have carried in the laboratory have shown that heparin can break up these aggregates and now I seek to characterize both crotamine-DNA and crotamine-GAG aggregates. Researchers have demonstrated that the internalization of these peptides is dependent on the size of the complex, signifying that different aggregate sizes have different transfection efficiencies. Therefore, the rate of endocytosis is dependent on the size of these aggregates and the optimum size will lead to better gene delivery. Because crotamine carries DNA into the cell in aggregated form, the characterization of these complexes is extremely important for understanding of this cell penetrating protein. The purpose of this research is to determine the conditions under which these aggregates form and the size of these aggregates, including the hydrodynamic radii.