Inhibition of autotransporter biogenesis by small molecules

Abstract
Disarming pathogens by targeting virulence factors is a promising alternative to classic antibiotics. Many virulence factors in Gram-negative bacteria are secreted via the autotransporter (AT) pathway, also known as Type 5 secretion. These factors are secreted with the assistance of two membrane-based protein complexes: Sec and Bam. To identify inhibitors of the AT pathway we used transcriptomics analysis to develop a fluorescence-based high-throughput assay that reports on the stress induced by the model AT hemoglobin protease (Hbp) when its secretion across the outer membrane is inhibited. Screening a library of 1600 fragments yielded the compound VUF15259 that provokes cell envelope stress and secretion inhibition of the ATs Hbp and Antigen-43. VUF15259 also impairs β-barrel folding activity of various outer membrane proteins. Furthermore, we found that mutants that are compromised in outer membrane protein biogenesis are more susceptible to VUF15259. Finally, VUF15259 induces the release of vesicles that appear to assemble in short chains. Taken together, VUF15259 is the first reported compound that inhibits AT secretion and our data are mostly consistent with VUF15259 interfering with the Bam-complex as potential mode of action. The validation of the presented assay incites its use to screen larger compound libraries with drug-like compounds. This article is protected by copyright. All rights reserved.

Citation
Steenhuis M, Abdallah A, de Munnik SM, Kuhne S, Sterk G, et al. (2019) Inhibition of autotransporter biogenesis by small molecules. Molecular Microbiology. Available: http://dx.doi.org/10.1111/mmi.14255.

Acknowledgements
We thank Hans Custers for synthetic efforts, and Iwan de Esch, Wilbert Bitter, Peter van Ulsen, AlexanderSpeer, Tanneke den Blaauwen, Corinne M. Ten Hagen-Jongman, Henry F. Vischer and Frank J. Bruggemanfor valuable input in the project and critical reading of the manuscript.

Publisher
Wiley

Journal
Molecular Microbiology

DOI
10.1111/mmi.14255

Additional Links
https://onlinelibrary.wiley.com/doi/abs/10.1111/mmi.14255

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