Abstract
The type VI secretion system (T6SS) delivers enzymatic effectors into target cells to destroy them. Cells of the same strain protect themselves against effectors with immunity proteins that specifically inhibit effectors. Here, we report the identification and characterization of a Tle3 phospholipase effector and its cognate immunity protein Tli3—an outer membrane lipoprotein from adherent-invasive Escherichia coli (AIEC). Enzymatic assays demonstrate that purified Tle3AIEC has a phospholipase A1, and not A2, activity and that its toxicity is neutralized by the cognate immunity protein Tli3AIEC. Tli3AIEC binds Tle3 in a 1:1 stoichiometric ratio. Tle3AIEC, Tli3AIEC and the Tle3AIEC-Tli3AIEC complex were purified and subjected to crystallization. The Tle3AIEC-Tli3AIEC complex structure could not be solved by SeMet phasing, but only by molecular replacement when using an AlphaFold2 prediction model. Tle3AIEC exhibits an α/β-hydrolase fold decorated by two protruding segments, including a N-terminus loop. Tli3AIEC displays a new fold of three stacked β-sheets and a protruding loop that inserts in Tle3AIECcatalytic crevice. We showed, experimentally, that Tle3AIEC interacts with the VgrG AIEC cargo protein and AlphaFold2 prediction of the VgrGAIEC-Tle3AIEC complex reveals a strong interaction between the VgrGAIEC C-terminus adaptor and Tle3AIEC N-terminal loop.
Funder
Aix-Marseille Université (AMU), the Centre National de la Recherche Scientifique
Agence Nationale de la Recherche
Subject
Inorganic Chemistry,Organic Chemistry,Physical and Theoretical Chemistry,Computer Science Applications,Spectroscopy,Molecular Biology,General Medicine,Catalysis
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