PMID:16954226

Citation	Cordero, CL, Kudryashov, DS, Reisler, E and Satchell, KJ (2006) The Actin cross-linking domain of the Vibrio cholerae RTX toxin directly catalyzes the covalent cross-linking of actin. J. Biol. Chem. 281:32366-74
Abstract	Vibrio cholerae is a Gram-negative bacterial pathogen that exports enterotoxins to alter host cells and to elicit diarrheal disease. Among the secreted toxins is the multifunctional RTX toxin, which causes cell rounding and actin depolymerization by covalently cross-linking actin monomers into dimers, trimers, and higher multimers. The region of the toxin responsible for cross-linking activity is the actin cross-linking domain (ACD). In this study, we further investigated the role of the ACD in the actin cross-linking reaction. We show that the RTX toxin cross-links actin independently of tissue transglutaminase, thus eliminating an indirect model of ACD activity. We demonstrate that a fusion protein of the ACD and the N-terminal portion of lethal factor from Bacillus anthracis (LF(N)ACD) has cross-linking activity in vivo and in crude cell extracts. Furthermore, we determined that LF(N)ACD directly catalyzes the formation of covalent linkages between actin molecules in vitro and that Mg(2+) and ATP are essential cofactors for the cross-linking reaction. In addition, G-actin is proposed as a cytoskeletal substrate of the RTX toxin in vivo. Future studies of the in vitro cross-linking reaction will facilitate characterization of the enzymatic properties of the ACD and contribute to our knowledge of the novel mechanism of covalent actin cross-linking.
Links	PubMed PMC2255562 Online version:10.1074/jbc.M605275200
Keywords	Actins/chemistry; Actins/metabolism; Acyltransferases/chemistry; Acyltransferases/genetics; Acyltransferases/metabolism; Bacterial Proteins/chemistry; Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Catalysis; Cell Extracts/chemistry; Cell Line, Tumor; Cross-Linking Reagents/chemistry; Cross-Linking Reagents/metabolism; Humans; Protein Structure, Tertiary; Vibrio cholerae/chemistry; Vibrio cholerae/genetics; Vibrio cholerae/metabolism; Vibrio cholerae/pathogenicity
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
VIBCH:Q9KS12	involved_in	GO:0018262: isopeptide cross-linking	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete
VIBCH:Q9KS12	involved_in	GO:0030042: actin filament depolymerization	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete
VIBCH:Q9KS12	enables	GO:0000287: magnesium ion binding	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete
VIBCH:Q9KS12	enables	GO:0005524: ATP binding	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete

References

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