Structural and Mechanistic Studies of Andropin, A Membrane-Selective Antimicrobial Peptide

Andropin is a 34 residue peptide, which originates from the testes of Drosophilia melanogaster and functions as an antimicrobial for the male reproductive tract. Andropin has activity against gram-positive bacteria, likely by selective disruption of the membrane wall, which causes cell lysis. The biological properties of this peptide have characteristics that are similar to both Cecropin and ALL38, but Andropin's structural properties and mechanistic details for membrane disruption have not yet been examined. Here, we examined Andropin's structural properties and its mechanism to selectively disrupt lipid membranes by using model membrane mimics of gram-positive, gram-negative, and mammalian cellular membranes. Turbidity measurements, as well as fluorescence leakage, FRET, and fluorescence anisotropy measurements were used to determine Andropin's effectiveness to disrupt these model membrane mimics. The secondary structure of the peptide was also probed with and without membrane environments using FTIR, CD, and NMR spectroscopies. Our preliminary results indicate that lipid head groups, particularly negatively charged species, drastically change Andropin's rate of membrane disruption. DPH anisotropy, combined with FRET measurements, indicates that Andropin functions either through a torodial pore or barrel-stave mechanism to pierce the membrane surface. Pore formation is supported by preliminary FTIR data that reveals a transition from a helix-turn-helix structure, when Andropin is free in solution, to a helix bundle upon interaction with negatively charged membranes. Ultimately, we plan to further characterize the high-resolution structure of Andropin and its membrane penetrating properties using an array of spectroscopic techniques. These results will help us understand how antimicrobial peptides can be designed to obtain properties for selective disruption of cellular membranes.