Interaction of theta-toxin (perfringolysin O), a cholesterol-binding cytolysin, with liposomal membranes: change in the aromatic side chains upon binding and insertion.
To understand the mechanism of membrane lysis by theta-toxin (perfringolysin O) from Clostridium perfringens, a cholesterol-binding, pore-forming cytolysin, we undertook a spectroscopic analysis of the structural changes that occur during the lytic process using lipid vesicles. In particular, the spectra were compared with those obtained using a modified theta-toxin, MC theta, that binds membrane cholesterol without forming oligomeric pores, thus bypassing the oligomerization step. The interaction of theta-toxin with liposomes composed of cholesterol and phosphatidylcholine but not with cholesterol-free liposomes caused a remarkable increase in the intensity of the tryptophan fluorescence emission spectra and ellipticity changes in the near- and far-UV CD peaks. A CD peak shift from 292 to 300 nm was specific for theta-toxin, suggesting oligomerization-specific changes occurring around tryptophan residues. Structural changes in the aromatic side chains were detected in the near-UV CD and fluorescence spectra upon MC theta-liposome interaction, although the far-UV CD spectra indicate that the beta-rich secondary structure of MC theta is well-conserved after membrane binding. Quenching of the intrinsic tryptophan fluorescence of MC theta by brominated lecithin/cholesterol liposomes suggests that theta-toxin inserts at least partly into membranes in the absence of oligomerization. These results indicate that regardless of oligomerization, the binding of theta-toxin to cholesterol induces partial membrane insertion and triggers conformational changes accompanied by aromatic side chain rearrangement with retention of secondary structure.(ABSTRACT TRUNCATED AT 250 WORDS)
Nakamura
M
M
Department of Enzyme Biochemistry and Membrane Biochemistry, Tokyo Metropolitan Institute of Gerontology, Japan.
Sekino
N
N
Iwamoto
M
M
Ohno-Iwashita
Y
Y
D001427:Bacterial Toxins / Q000737:chemistry* / Q000378:metabolism; D002942:Circular Dichroism; D003016:Clostridium perfringens / Q000737:chemistry / Q000472:pathogenicity; D004910:Erythrocyte Membrane / Q000737:chemistry; D006460:Hemolysin Proteins; D066298:In Vitro Techniques; D008081:Liposomes / Q000737:chemistry*; D008567:Membranes, Artificial; D010713:Phosphatidylcholines / Q000737:chemistry; D017433:Protein Structure, Secondary; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D014364:Tryptophan / Q000737:chemistry
D001427:Bacterial Toxins; D006460:Hemolysin Proteins; D008081:Liposomes; D008567:Membranes, Artificial; D010713:Phosphatidylcholines; C020452:Clostridium perfringens theta-toxin; D014364:Tryptophan