Structure of a Rhs effector clade domain identifies new mechanistic insights into type VI secretion system toxin delivery.

Hayes, B. K.; Harper, M.; Venugopal, H.; Lewis, J. M.; Wright, A.; Steele, J. R.; Steer, D.; Schittenhelm, R. B.; Boyce, J. D.; McGowan, S.

doi:10.1101/2023.11.18.567679

bioRxiv · 10.1101/2023.11.18.567679

Structure of a Rhs effector clade domain identifies new mechanistic insights into type VI secretion system toxin delivery.

Hayes, B. K.·Harper, M.·Venugopal, H.·Lewis, J. M.·Wright, A.·Steele, J. R.·Steer, D.·Schittenhelm, R. B.·Boyce, J. D.·McGowan, S.

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Abstract

The type VI secretion system (T6SS) is a molecular machine utilised by many Gram-negative bacteria to deliver antibacterial toxins directly into adjacent bacteria, often providing a competitive advantage. Rearrangement hotspot (Rhs) effectors are known T6SS cargo effectors, but the precise delivery and activation of these toxins is poorly defined. We present the structure of a novel T6SS Rhs effector (Tse15) from the multidrug resistant nosocomial pathogen Acinetobacter baumannii. Tse15 forms a triple layered y-cocoon Rhs domain with an N-terminal -helical clade domain and an unfolded C-terminal toxin domain located entirely inside the Rhs cage. We identify that Tse15 is cleaved into three domains, through two independent auto-cleavage events involving aspartyl protease activity for toxin self-cleavage and a nucleophilic glutamic acid for the cleavage of the N-terminal clade domain. Proteomic analyses showed that the N-terminal clade and toxin domains, but not the y-strand rich Rhs cage, are delivered outside of the cell, suggesting a novel mechanism for Rhs toxin delivery and activation. Our findings suggest that this delivery mechanism requires an interaction between the N-terminal clade and toxin domains, with the clade domain acting as the internal chaperone to mediate tethering of the toxin to the T6SS machinery. Conservation of the clade domain in other Gram-negative bacteria suggest this may be a common mechanism for T6SS toxin delivery.

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Hayes, B. K., Harper, M., Venugopal, H., Lewis, J. M., Wright, A., Steele, J. R., Steer, D., Schittenhelm, R. B., Boyce, J. D., McGowan, S.. 2023-11-18. Structure of a Rhs effector clade domain identifies new mechanistic insights into type VI secretion system toxin delivery.. https://doi.org/10.1101/2023.11.18.567679

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