Selective Lysis of Bacteria but Not Mammalian Cells by Diastereomers of Melittin: Structure−Function Study

Abstract

Studies on lipid-peptide interactions of cytolytic polypeptides tend to emphasize the importance of the amphipathic alpha-helical structure for their cytolytic activity. In this study, diasetereomers of the bee venom melittin (26 a.a.), a non-cell-selective cytolysin, were synthesized and investigated for their structure and cytolytic activity toward bacteria and mammalian cells. Similarly to the findings with the diastereomers of the less cytolytic peptide pardaxin (33 a.a.) (Shai & Oren. 1996), the melittin diastereomer, lest their alpha-helical structure, which abrogated their hemolytic activity toward human erythrocytes. However, they retained their antibacterial activity and completely lysed both Gram-positive and Gram-negative bacteria, as revealed by transmission electron microscopy. To understand the molecular mechanism underlying this selectivity, binding experiments utilizing the intrinsic tryptophan of melittin, tryptophan quenching experiments using brominated phospholipids, and membrane destabilization studies were done. The data revealed that the melittin diastereomers bound to and destabilized only negatively-charged phospholipid vesicles, in contrast to native melittin, which binds strongly to both negatively-charged and zwitterionic phospholipids. However, the partition coefficient, the depth of penetration into the membrane, and the membrane-permeating activity of the diastereomers with negatively-charged phospholipids were similar to those obtained with melittin. The results obtained do not support the formation of transmembrane pores as the mode of action of the diastereomers, but rather suggest that these peptides bind to the surface of the bacterial membrane, cover it in a "carpet-like" manner, and dissolve it like a detergent. The results presented here together with those obtained with the cytolytic peptide pardaxin suggest that the combination of hydrophobicity and net positive charge may be sufficient in the design of potent diastereomers of antibacterial polypeptides for the treatment of infectious diseases.