Structure of the claudin-binding domain of Clostridium perfringens enterotoxin.
Clostridium perfringens enterotoxin is a common cause of food-borne and antibiotic-associated diarrhea. The toxin's receptors on intestinal epithelial cells include claudin-3 and -4, members of a large family of tight junction proteins. Toxin-induced cytolytic pore formation requires residues in the NH(2)-terminal half, whereas residues near the COOH terminus are required for binding to claudins. The claudin-binding COOH-terminal domain is not toxic and is currently under investigation as a potential drug absorption enhancer. Because claudin-4 is overexpressed on some human cancers, the toxin is also being investigated for targeting chemotherapy. Our aim was to solve the structure of the claudin-binding domain to advance its therapeutic applications. The structure of a 14-kDa fragment containing residues 194 to the native COOH terminus at position 319 was solved by x-ray diffraction to a resolution of 1.75A. The structure is a nine-strand beta sandwich with previously unappreciated similarity to the receptor-binding domains of several other toxins of spore-forming bacteria, including the collagen-binding domain of ColG from Clostridium histolyticum and the large Cry family of toxins (including Cry4Ba) of Bacillus thuringiensis. Correlations with previous studies suggest that the claudin-4 binding site is on a large surface loop between strands beta8 and beta9 or includes these strands. The sequence that was crystallized (residues 194-319) binds to purified human claudin-4 with a 1:1 stoichiometry and affinity in the submicromolar range similar to that observed for binding of native toxin to cells. Our results provide a structural framework to advance therapeutic applications of the toxin and suggest a common ancestor for several receptor-binding domains of bacterial toxins.
Van Itallie
Christina M
CM
Department of Medicine, Chapel Hill, North Carolina 27599.
Betts
Laurie
L
Department of Pharmacology, Chapel Hill, North Carolina 27599.
Smedley
James G
JG
Department of Molecular Genetics and Biochemistry, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261.
McClane
Bruce A
BA
Department of Molecular Genetics and Biochemistry, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261.
Anderson
James M
JM
Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, North Carolina 27599. Electronic address: jandersn@med.unc.edu.
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D062506:Claudin-4; D003016:Clostridium perfringens / Q000378:metabolism*; D004768:Enterotoxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D040681:Structural Homology, Protein; D014961:X-Ray Diffraction
D001426:Bacterial Proteins; C568157:CLDN4 protein, human; D062506:Claudin-4; D004768:Enterotoxins; D008565:Membrane Proteins; C035598:enterotoxin, Clostridium