PMID:9426125

Citation	Guillouard, I, Alzari, PM, Saliou, B and Cole, ST (1997) The carboxy-terminal C2-like domain of the alpha-toxin from Clostridium perfringens mediates calcium-dependent membrane recognition. Mol. Microbiol. 26:867-76
Abstract	The lethal, cytolytic alpha-toxin (phospholipase C) of Clostridium perfringens consists of two distinct modules: the larger N-terminal domain catalyses phospholipid hydrolysis, and its activity is potentiated by a smaller C-terminal domain. Calcium ions are essential for the binding of alpha-toxin to lipid films. Sixteen alpha-toxin variants with single amino acid substitutions in the C-terminal region were obtained using site-directed mutagenesis and T7 expression technology. Five of these variants showed reduced phospholipase C activity and were considerably less active than native alpha-toxin under calcium-limiting conditions. Replacement of Thr-272 by Pro diminished phospholipase C activity, severely affected haemolysis and platelet aggregation and perturbed a surface-exposed conformational epitope. The results of sequence comparisons and molecular modelling indicate that the C-terminal region probably belongs to the growing family of C2 beta-barrel domains, which are often involved in membrane interactions, and that the functionally important substitutions are clustered at one extremity of the domain. The combined findings suggest that the C-terminal region of alpha-toxin mediates interactions with membrane phospholipids in a calcium-dependent manner. Mutations to this domain may account for the natural lack of toxicity of the alpha-toxin homologue, phospholipase C of Clostridium bifermentans.
Links	PubMed
Keywords	Amino Acid Sequence; Animals; Bacterial Toxins/genetics; Bacterial Toxins/metabolism; Binding Sites; Calcium/metabolism; Calcium/pharmacology; Calcium-Binding Proteins; Cell Membrane/metabolism; Clostridium perfringens/enzymology; Clostridium perfringens/genetics; Culture Media/pharmacology; Humans; Molecular Sequence Data; Proline/genetics; Proline/metabolism; Protein Structure, Tertiary; Sequence Homology, Amino Acid; Sheep; Threonine/genetics; Threonine/metabolism; Type C Phospholipases/genetics; Type C Phospholipases/metabolism; Tyrosine/genetics; Tyrosine/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
CLOPE:PHLC1		GO:0050429: calcium-dependent phospholipase C activity	ECO:0000314:		F		Table 1: alpha toxin variants decrease phospholipase C activation when Tyr-275 and Tyr-277 are changed in recombinant toxin	complete CACAO 11313
CLOPE:PHLC1		GO:0007202: activation of phospholipase C activity	ECO:0000314:		P		Figure 2 shows direct correlation between calcium concentrations and its role on phospholipase C activation.	complete CACAO 11314

References

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