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Monitoring the Assembly of a Secreted Bacterial Virulence Factor Using Site-specific Crosslinking
11:33

Monitoring the Assembly of a Secreted Bacterial Virulence Factor Using Site-specific Crosslinking

Published on: December 17, 2013

Mechanisms of assembly and cellular interactions for the bacterial genotoxin CDT

Dragana Nesic1, C Erec Stebbins

  • 1Laboratory of Structural Microbiology, The Rockefeller University, New York, New York, USA.

Plos Pathogens
|November 24, 2005
PubMed

Insights

Cytolethal distending toxin (CDT) uses CdtA and CdtC subunits to bind host cells, causing DNA damage. Structural analysis reveals key binding elements crucial for CDT

Area of Science:

  • Bacterial pathogenesis
  • Molecular toxicology
  • Structural biology

Background:

  • Bacterial pathogens utilize cytolethal distending toxin (CDT) to induce host cell DNA damage, cell cycle arrest, and apoptosis.
  • CDT is a tripartite holotoxin comprising a DNase I family nuclease (CdtB) and two ricin-like lectin domains (CdtA and CdtC).

Purpose of the Study:

  • To investigate the structural elements of CdtA and CdtC subunits essential for CDT holotoxin assembly, cell surface binding, and toxic activity.
  • To elucidate the functional roles of specific binding sites within the CDT holotoxin.

Main Methods:

  • Structure-based mutagenesis was employed to modify key residues in CdtA and CdtC.
  • Biochemical assays were used to assess toxin assembly and stability.
  • Cellular toxicity assays and cell surface binding assays (e.g., on HeLa cells) were performed to evaluate functional consequences.

Main Results:

  • N- and C-terminal nonglobular polypeptides of CdtA and CdtC significantly contribute to holotoxin stability and activity through interactions with CdtB.
  • An aromatic cluster in CdtA and a groove at the CdtA-CdtC interface are identified as critical binding elements.
  • Mutations in these binding elements impair HeLa cell binding and subsequent cellular intoxication, demonstrating a strong correlation between binding and activity.

Conclusions:

  • The study establishes structure-based hypotheses for the assembly and function of the CDT toxin family.
  • Specific structural features of CdtA and CdtC are critical for CDT-mediated host cell DNA damage and pathogenesis.

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