BACTERICIDAL ACTIVITY OF LYS49 AND ASP49 MYOTOXIC PHOSPHOLIPASES A(2)FROM BOTHROPS-ASPER SNAKE-VENOM - SYNTHETIC LYS49 MYOTOXIN II-(115-129)-PEPTIDE IDENTIFIES ITS BACTERICIDAL REGION

Mammalian group-II phospholipases A(2) (PLA(2)) of inflammatory fluids display bactericidal properties, which are dependent on their enzymat ic activity. This study shows that myotoxins II (Lys49) and III (Asp49 ), two group-II PLA(2) isoforms from the venom of Bothrops asper, are lethal to a broad spectrum of bacteria. Since the catalytically inacti ve Lys49 myotoxin II isoform has similar bactericidal effects to its c atalytically active Asp49 counterpart, a bactericidal mechanism that i s independent of an intrinsic PLA(2) activity is demonstrated. Moreove r, a synthetic 13-residue peptide of myotoxin II, comprising residues 115-129 (common numbering system) near the C-terminal loop, reproduced the bactericidal effect of the intact protein. Following exposure to the peptide or the protein, accelerated uptake of the hydrophobic prob e N-phenyl-N-naphthylamine was observed in susceptible but not in resi stant bacteria, indicating that the lethal effect was initiated on the bacterial membrane. The outer membrane, isolated lipopolysaccharide ( LPS), and lipid A of susceptible bacteria showed higher binding to the myotoxin II-(115-129)-peptide than the corresponding moieties of resi stant strains. Bacterial LPS chimeras indicated that LPS is a relevant target for myotoxin II-(115-129)-peptide. When heterologous LPS of th e resistant strain was present in the context of susceptible bacteria, the chimera became resistant, and vice versa. Myotoxin II represents a group-II PLA(2) with a direct bactericidal effect that is independen t of an intrinsic enzymatic activity, but adscribed to the presence of a short cluster of basic/hydrophobic amino acids near its C-terminal loop.