Structure-function analysis of inositol hexakisphosphate-induced autoprocessing in Clostridium difficile toxin A.

Pruitt RN, Chagot B, Cover M, Chazin WJ, Spiller B, Lacy DB
J Biol Chem. 2009 284 (33): 21934-40

PMID: 19553670 · PMCID: PMC2755918 · DOI:10.1074/jbc.M109.018929

The action of Clostridium difficile toxins A and B depends on inactivation of host small G-proteins by glucosylation. Cellular inositol hexakisphosphate (InsP6) induces an autocatalytic cleavage of the toxins, releasing an N-terminal glucosyltransferase domain into the host cell cytosol. We have defined the cysteine protease domain (CPD) responsible for autoprocessing within toxin A (TcdA) and report the 1.6 A x-ray crystal structure of the domain bound to InsP6. InsP6 is bound in a highly basic pocket that is separated from an unusual active site by a beta-flap structure. Functional studies confirm an intramolecular mechanism of cleavage and highlight specific residues required for InsP6-induced TcdA processing. Analysis of the structural and functional data in the context of sequences from similar and diverse origins highlights a C-terminal extension and a pi-cation interaction within the beta-flap that appear to be unique among the large clostridial cytotoxins.

MeSH Terms (16)

Bacterial Toxins Catalytic Domain Cations Clostridium difficile Crystallography, X-Ray Enterotoxins Magnetic Resonance Spectroscopy Models, Biological Models, Molecular Molecular Conformation Phytic Acid Point Mutation Protein Structure, Secondary Protein Structure, Tertiary Spectrophotometry Time Factors

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