Membrane Interactions of Streptococcus agalactiae's CAMP factor

Abstract

CAMP factor is an extracellular pore-forming toxin secreted by the group B streptococci Streptococcus agalactiae. In conjunction with the action of sphingomyelinase secreted by Staphylococcus aureus, which converts membrane sphingomyline to ceramide, CAMP factor kills susceptible cells by creating holes in them. Since the monomeric or oligomeric structure of CAMP factor is not yet known, no studies on the membrane-penetrating domain of this toxin have been done. In the present study, the interaction of a putative hydrophobic domain between residues T90 and V115 with the target membrane was examined by cysteine-scanning mutagenesis and site-selective fluorescent labeling. The combination of steady state and lifetime fluorescence measurements and collisional quenching experiments with nitroxide labeled fatty acids indicate that residues from T90 to V115 contact the membrane upon binding and oligomerization of CAMP factor on cell membranes. More importantly, all these individual assays indicate that the residues from N104C to F109C insert superficially into the membrane with a β-sheet conformation.