The influences of the antibacterial magainin 2 and PGLa from the African clawed frog (Xenopus laevis) and the hemolytic bee venom melittin on Escherichia coli as the target cell were studied by atomic force microscopy (AFM). Nanometer-scale images of the effects of the peptides on this gram-negative bacterium's cell envelope were obtained in situ without the use of fixing agents. These high-resolution AFM images of the surviving and intact target cells before and after peptide treatment showed distinct changes in cell envelope morphology as a consequence of peptide action. Although all three peptides are lytic to E. coli, it is clear from this AFM study that each peptide causes distinct morphological changes in the outer membrane and in some cases the inner membrane, probably as a consequence of different mechanisms of action.Bacterial resistance to conventional antibiotics has become a major problem worldwide, and certain strains of bacteria are already resistant to all available drugs (3, 29). The development of a new family of antibiotics is therefore an important research topic. Antimicrobial peptides are considered good candidates for the next group of antibiotics because their proposed mode of action is different from those of conventional antibiotics (12,25). To advance antimicrobial peptides to the status of a new group of antibiotics, it is important to understand the mechanisms of action of these agents and the reason for their selectivity against microbes. In this study we compared the actions of three different peptides, namely, melittin, the toxic and hemolytic 26-residue peptide from the European honeybee Apis mellifera (6, 11), and two antibacterial peptides from the African clawed frog Xenopus laevis, an amide analogue of the 23-residue magainin 2 (32, 33) and the 21-residue PGLa (28), on the membrane of Escherichia coli. All three peptides are cationic ␣-helical antimicrobial peptides with markedly different sequential distributions of polar and nonpolar amino acids. However, all three peptides are amphipathic and highly membranolytic, with PGLa and magainin 2 having a better selectivity, via electrostatic discrimination, toward prokaryotic cells (13, 22) than the hemolytic melittin (which is a nonselective lytic peptide).Model membrane studies have provided a good understanding of antimicrobial peptides and their mechanisms of action. Different models for the interaction of a peptide with the bacterial membrane have been proposed: the barrel stave model (2), the toroidal pore or wormhole model (5, 31), the carpet model (23), and the peptide aggregate model (19,30). Most antimicrobial peptides are thought to follow the toroidal pore model and/or carpet model and not the barrel stave model, in which the peptides in the pore need to span the membrane without deformation of the lipid bilayer. Peptides that form toroidal pores remain associated with the phospholipid head groups and insert perpendicularly to the membrane plane by causing the lipid bilayer to fold back on itself. It has been proposed ...

Pseudomonas strain CF600 is able to utilize phenol and 3,4-dimethylphenol as sole carbon and energy source. We demonstrate that growth on these substrates is by virtue of plasmid-encoded phenol hydroxylase and a meta-cleavage pathway. Screening of a genomic bank, with DNA from the previously cloned catechol2,3-dioxygenase gene of the TOL plasmid pWW0, was used in the identification of a clone which could complement a phenol-hydroxylase-deficient transposon insertion mutant. Deletion mapping and polypeptide production analysis identified a 1.2 kb region of DNA encoding a 39-5 kDa polypeptide which mediated this complementation. Enzyme activities and growth properties of Pseudomonas strains harbouring this fragment on a broad-host-range expression vector indicate that phenol hydroxylase is a multicomponent enzyme containing the 39.5 kDa polypeptide as one component.