Designed Ankyrin Repeat Proteins provide insights into the structure and function of CagI and are potent inhibitors of CagA translocation by the Helicobacter pylori type IV secretion system.

Autor: Blanc M; UMR 5086 Molecular Microbiology and Structural Biochemistry CNRS-Université de Lyon, Institut de Biologie et Chimie des Protéines, Lyon, France., Lettl C; Max von Pettenkofer Institute of Hygiene and Medical Microbiology, Faculty of Medicine, LMU Munich, Munich, Germany., Guérin J; UMR 5086 Molecular Microbiology and Structural Biochemistry CNRS-Université de Lyon, Institut de Biologie et Chimie des Protéines, Lyon, France., Vieille A; UMR 5086 Molecular Microbiology and Structural Biochemistry CNRS-Université de Lyon, Institut de Biologie et Chimie des Protéines, Lyon, France., Furler S; Department of Biochemistry, University of Zurich, Zurich, Switzerland., Briand-Schumacher S; Department of Biochemistry, University of Zurich, Zurich, Switzerland., Dreier B; Department of Biochemistry, University of Zurich, Zurich, Switzerland., Bergé C; UMR 5086 Molecular Microbiology and Structural Biochemistry CNRS-Université de Lyon, Institut de Biologie et Chimie des Protéines, Lyon, France., Plückthun A; Department of Biochemistry, University of Zurich, Zurich, Switzerland., Vadon-Le Goff S; University of Lyon, CNRS UMR5305, Tissue Biology and Therapeutic Engineering Laboratory (LBTI), Lyon, France., Fronzes R; European Institute of Chemistry and Biology, CNRS UMR 5234 Microbiologie Fondamentale et Pathogénicité, Univ. Bordeaux, Pessac, France., Rousselle P; University of Lyon, CNRS UMR5305, Tissue Biology and Therapeutic Engineering Laboratory (LBTI), Lyon, France., Fischer W; Max von Pettenkofer Institute of Hygiene and Medical Microbiology, Faculty of Medicine, LMU Munich, Munich, Germany., Terradot L; UMR 5086 Molecular Microbiology and Structural Biochemistry CNRS-Université de Lyon, Institut de Biologie et Chimie des Protéines, Lyon, France.
Jazyk: angličtina
Zdroj: PLoS pathogens [PLoS Pathog] 2023 May 08; Vol. 19 (5), pp. e1011368. Date of Electronic Publication: 2023 May 08 (Print Publication: 2023).
DOI: 10.1371/journal.ppat.1011368
Abstrakt: The bacterial human pathogen Helicobacter pylori produces a type IV secretion system (cagT4SS) to inject the oncoprotein CagA into gastric cells. The cagT4SS external pilus mediates attachment of the apparatus to the target cell and the delivery of CagA. While the composition of the pilus is unclear, CagI is present at the surface of the bacterium and required for pilus formation. Here, we have investigated the properties of CagI by an integrative structural biology approach. Using Alpha Fold 2 and Small Angle X-ray scattering, it was found that CagI forms elongated dimers mediated by rod-shape N-terminal domains (CagIN) prolonged by globular C-terminal domains (CagIC). Three Designed Ankyrin Repeat Proteins (DARPins) K2, K5 and K8 selected against CagI interacted with CagIC with subnanomolar affinities. The crystal structures of the CagI:K2 and CagI:K5 complexes were solved and identified the interfaces between the molecules, thereby providing a structural explanation for the difference in affinity between the two binders. Purified CagI and CagIC were found to interact with adenocarcinoma gastric (AGS) cells, induced cell spreading and the interaction was inhibited by K2. The same DARPin inhibited CagA translocation by up to 65% in AGS cells while inhibition levels were 40% and 30% with K8 and K5, respectively. Our study suggests that CagIC plays a key role in cagT4SS-mediated CagA translocation and that DARPins targeting CagI represent potent inhibitors of the cagT4SS, a crucial risk factor for gastric cancer development.
Competing Interests: The authors have declared that no competing interests exist.
(Copyright: © 2023 Blanc et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)
Databáze: MEDLINE
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