Characterization of Streptococcus agalactiae CAMP factor as a pore-forming toxin.
A recombinant form of CAMP factor of Streptococcus agalactiae has been expressed as glutathione S-transferase-CAMP fusion protein in Escherichia coli. After thrombin cleavage of the fusion protein, the recombinant CAMP factor exhibited hemolytic activity comparable with that of the native form. Osmotic protection experiments with polyethylene glycols show that CAMP factor forms discrete transmembrane pores with a diameter upward of 1.6 nm on susceptible membranes; electron microscopy reveals circular membrane lesions of heterogeneous size, up to 12-15 nm in diameter. Liposome permeabilization studies show that pore formation is a highly cooperative process, which suggests that it involves the oligomerization of CAMP factor. Chemical cross-linking experiments also support an oligomeric mode of action.
Lang
Shenhui
S
Department of Chemistry, University of Waterloo, Waterloo, Ontario N2L 3G1, Canada.
Palmer
Michael
M
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry*; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D002942:Circular Dichroism; D003001:Cloning, Molecular; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D005452:Fluoresceins / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006460:Hemolysin Proteins / Q000737:chemistry; D008081:Liposomes / Q000378:metabolism; D008854:Microscopy, Electron; D010957:Plasmids / Q000378:metabolism; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D012756:Sheep; D013292:Streptococcus agalactiae / Q000378:metabolism*; D013997:Time Factors; D014118:Toxins, Biological / Q000737:chemistry*
D001426:Bacterial Proteins; C039092:CAMP protein, Streptococcus; D003432:Cross-Linking Reagents; D005452:Fluoresceins; D006460:Hemolysin Proteins; D008081:Liposomes; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014118:Toxins, Biological; D005982:Glutathione Transferase; C007740:fluorexon