Membrane activity of the peptide antibiotic clavanin and the importance ofits glycine residues

The peptide antibiotic clavanin A (VFQFLGKIIHHVGNFVHGFSHVF-NH2) is rich in histidine and glycine residues. In this study the antimicrobial activity an d membrane activity of wild-type clavanin A and seven Gly --> Ala mutants t hereof were investigated. Clavanin A effectively killed the test microorgan ism Micrococcus flavus and permeabilized its cytoplasmic membrane in the mi cromolar concentration range, suggesting that the membrane is the target fo r this molecule. Consistent with this suggestion, it was observed that clav anin A efficiently inserted into different phospholipid monolayers mainly v ia hydrophobic interactions. Bilayer permeabilization was observed for both low and high molecular mass fluorophores enclosed in unilamellar vesicles and occurred at the same concentration as the antimicrobial activity. It is therefore suggested that the loss of barrier function does not involve spe cific receptors in the target membrane. Circular dichroism spectroscopy ind icated that under membrane mimicking conditions a random coil --> helical t ransition was induced for all clavanin derivatives tested. Observed differe nces in peptide-membrane interaction and biological activity between the va rious clavanin derivatives demonstrated the functional importance of Gly at the positions 6 and 13. These two glycines may act as flexible hinges that facilitate the hydrophobic N-terminal end of clavanin to deeply insert int o the bilayer. On the contrary, no such role is evident for Gly 18, as its substitution by Ala actually stimulated membrane interaction and biological activity. This study suggests that the combined hydrophobicity, overall st ate of charge, and conformational flexibility of the peptide determine the (membrane) activity of clavanin A and its Gly --> Ala mutants.