MODULATION OF STRUCTURE AND ANTIBACTERIAL AND HEMOLYTIC-ACTIVITY BY RING SIZE IN CYCLIC GRAMICIDIN-S ANALOGS

We have evaluated the effect of ring size of gramicidin S analogs on s econdary structure, lipid binding, lipid disruption, antibacterial and hemolytic activity, Cyclic analogs with ring sizes ranging from 4 to 14 residues were designed to maintain the amphipathic character as fou nd in gramicidin S and synthesized by solid phase peptide synthesis. T he secondary structure of these peptides showed a definite periodicity in beta-sheet content, with rings containing 6, 10, and 14 residues e xhibiting beta-sheet structure, and rings containing 8 or 12 residues being largely disordered, Peptides containing 4 or 6 residues did not bind lipopolysaccharide, whereas longer peptides showed a trend of inc reasing binding affinity for Lipopolysaccharide with increasing length , Destabilization of Escherichia coli outer membranes was only observe d in peptides containing 10 or more residues, Peptides containing fewe r than 10 residues were completely inactive and exhibited no hemolytic activity, The 10-residue peptide showed an activity profile similar t o that of gramicidin S itself, with activity against Gram-positive and Gram negative microorganisms as well as yeast, but also showed high h emolytic activity, Differential activities were obtained by increasing the size of the ring to either 12 or 14 residues, The 14-residue pept ide showed no antibiotic activity but exhibited increased hemolytic ac tivity, The 12-residue peptide lost activity against Gram-positive bac teria, retained activity against Gram-negative microorganisms and yeas t, but displayed decreased hemolytic activity, Biological activities i n the 12-residue peptide were optimized by a series of substitutions i n residues comprising both hydrophobic and basic sites resulting in a peptide that exhibited activities comparable with gramicidin S against Gram-negative microorganisms and yeast but with substantially lower h emolytic activity, Compared with gramicidin S, the best analog showed a 10-fold improvement in antibiotic specificity for Gram-negative micr oorganisms and a 7-fold improvement in specificity for yeast over huma n erythrocytes as determined by a therapeutic index. These results ind icate that it is possible to modulate structure and activities of cycl ic gramicidin S analogs by varying ring sizes and further show the pot ential for developing clinically useful antibiotics based on gramicidi n S.