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Structural Basis of Subtilase Cytotoxin SubAB Assembly

2013; Elsevier BV; Volume: 288; Issue: 38 Linguagem: Inglês

10.1074/jbc.m113.462622

1083-351X

Jérôme Le Nours, Adrienne W. Paton, Emma Byres, Sally Troy, Brock P. Herdman, Matthew D. Johnson, James C. Paton, Jamie Rossjohn, Travis Beddoe,

Bacterial Genetics and Biotechnology

Pathogenic strains of Escherichia coli produce a number of toxins that belong to the AB5 toxin family, which comprise a catalytic A-subunit that induces cellular dysfunction and a B-pentamer that recognizes host glycans. Although the molecular actions of many of the individual subunits of AB5 toxins are well understood, how they self-associate and the effect of this association on cytotoxicity are poorly understood. Here we have solved the structure of the holo-SubAB toxin that, in contrast to other AB5 toxins whose molecular targets are located in the cytosol, cleaves the endoplasmic reticulum chaperone BiP. SubA interacts with SubB in a similar manner to other AB5 toxins via the A2 helix and a conserved disulfide bond that joins the A1 domain with the A2 helix. The structure revealed that the active site of SubA is not occluded by the B-pentamer, and the B-pentamer does not enhance or inhibit the activity of SubA. Structure-based sequence comparisons with other AB5 toxin family members, combined with extensive mutagenesis studies on SubB, show how the hydrophobic patch on top of the B-pentamer plays a dominant role in binding the A-subunit. The structure of SubAB and the accompanying functional characterization of various mutants of SubAB provide a framework for understanding the important role of the B-pentamer in the assembly and the intracellular trafficking of this AB5 toxin.Background: AB5 toxins consist of a pentameric B-subunit and a catalytic A-subunit.Results: Crystallographic data, dissociation, and intracellular trafficking of SubAB toxin are reported.Conclusion: SubAB architecture is similar to other AB5 toxins, whereas the B-pentamer plays an important role in assembly and intracellular trafficking.Significance: The conserved hydrophobic ring in the B-pentamer supports the view that A- and B-subunits have evolved independently. Pathogenic strains of Escherichia coli produce a number of toxins that belong to the AB5 toxin family, which comprise a catalytic A-subunit that induces cellular dysfunction and a B-pentamer that recognizes host glycans. Although the molecular actions of many of the individual subunits of AB5 toxins are well understood, how they self-associate and the effect of this association on cytotoxicity are poorly understood. Here we have solved the structure of the holo-SubAB toxin that, in contrast to other AB5 toxins whose molecular targets are located in the cytosol, cleaves the endoplasmic reticulum chaperone BiP. SubA interacts with SubB in a similar manner to other AB5 toxins via the A2 helix and a conserved disulfide bond that joins the A1 domain with the A2 helix. The structure revealed that the active site of SubA is not occluded by the B-pentamer, and the B-pentamer does not enhance or inhibit the activity of SubA. Structure-based sequence comparisons with other AB5 toxin family members, combined with extensive mutagenesis studies on SubB, show how the hydrophobic patch on top of the B-pentamer plays a dominant role in binding the A-subunit. The structure of SubAB and the accompanying functional characterization of various mutants of SubAB provide a framework for understanding the important role of the B-pentamer in the assembly and the intracellular trafficking of this AB5 toxin. Background: AB5 toxins consist of a pentameric B-subunit and a catalytic A-subunit. Results: Crystallographic data, dissociation, and intracellular trafficking of SubAB toxin are reported. Conclusion: SubAB architecture is similar to other AB5 toxins, whereas the B-pentamer plays an important role in assembly and intracellular trafficking. Significance: The conserved hydrophobic ring in the B-pentamer supports the view that A- and B-subunits have evolved independently.