Interaction of the cyclic antimicrobial cationic peptide bactenecin with the outer and cytoplasmic membrane

Bactenecin, a 12-amino acid cationic antimicrobial peptide from bovine neut rophils, has two cysteine residues, which form one disulfide bond, making i t a cyclic molecule. To study the importance of the disulfide bond, a linea r derivative Bac2S was made and the reduced form (linear bactenecin) was al so included in this study. Circular dichroism spectroscopy showed that bact enecin existed as a type I p-turn structure regardless of its environment, while the reduced form and linear bactenecin adopted different conformation s according to the lipophilicity of the environment. Bactenecin was more ac tive against the Gram-negative wild type bacteria Escherichia coli, Pseudom onas aeruginosa, and Salmonella typhimurium than its linear derivative and reduced form, while all three peptides were equally active against the oute r membrane barrier-defective mutants of the first two bacteria. Only the tw o linear peptides showed activity against the Gram-positive bacteria Staphy lococcus epidermidis and Enterococcus facaelis, Bactenecin interacted well with the outer membrane and its higher affinity for E. coli UB1005 lipopoly saccharide and improved ability to permeabilize the outer membrane seemed t o account for its better antimicrobial activity against Gram-negative bacte ria. The interaction of bactenecin with the cytoplasmic membrane was determ ined by its ability to dissipate the membrane potential by using the fluore scence probe 3,3-dipropylthiacarbocyanine and an outer membrane barrier-def ective mutant E, coli DC2, It was shown that the linear derivative and redu ced form were able to dissipate the membrane potential at much lower concen trations than bactenecin despite the similar minimal inhibitory concentrati ons of all three against this barrier-defective mutant.