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Identification of small molecules blocking the Pseudomonas aeruginosa type III secretion system protein PcrV
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Science and Technology, Department of Chemistry.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Pseudomonas aeruginosa is an opportunistic bacterial pathogen that employs the type III secretion system (T3SS) during acute phase of infection to translocate cytotoxins into the host cell cytoplasm. PcrV, known as the V-antigen, is a T3S protein that facilitates the integration of the translocators into the eukaryotic cell membrane. In this study, we report surface plasmon resonance screening to identify small molecule binders of PcrV. A follow up primary structure-activity relationship analysis resulted in anti-virulence PcrV binders that protect macrophages in a  P. aeruginosa cell-based infection assay.

Keywords [en]
Pseudomonas aeruginosa, type III secretion, PcrV, surface plasmon resonance, screening, small molecules, macrophages
National Category
Natural Sciences
Research subject
Organic Chemistry; biology
Identifiers
URN: urn:nbn:se:umu:diva-150969OAI: oai:DiVA.org:umu-150969DiVA, id: diva2:1240416
Funder
Swedish Foundation for Strategic Research , SB12-0022Available from: 2018-08-21 Created: 2018-08-21 Last updated: 2018-08-21
In thesis
1. Towards novel antibacterials: Synthesis and identification of natural product inspired inhibitors of Chlamydia trachomatis and development of chemical probes targeting virulence of Pseudomonas aeruginosa
Open this publication in new window or tab >>Towards novel antibacterials: Synthesis and identification of natural product inspired inhibitors of Chlamydia trachomatis and development of chemical probes targeting virulence of Pseudomonas aeruginosa
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Antibiotic resistance has evolved significantly to become one of the serious threats to public health today. Yet, the pipeline of new antibiotics is drying up and is lagging behind the challenging needs. As a contribution to this recurrent need for novel antibacterials, we applied multidisciplinary strategies to identify small-molecule antibacterials against Chlamydia trachomatis and antivirulence agents against Pseudomonas aeruginosa infections. These strategies included:

1. Synthesis of a focused compounds library inspired by natural product scaffolds followed by phenotypic screening against Chlamydia trachomatis. (Paper I)

(-)-Hopeaphenol is a polyphenol natural product that was identified as an antivirulence agent against Y. pseudotuberculosis and P. aeruginosa. Hopeaphenol core scaffold, 2,3-diaryl-2,3-dihydrobenzofuran, is ubiquitous in polyphenolic phytochemicals. In this thesis, a focused library of forty-eight compounds was synthesized based on 2,3-diarylbenzofuran and 2,3-diaryl-2,3- dihydrobenzofuran. The library was then explored for antibacterial properties in a number of screening assays and resulted in five novel antichlamydial compounds with inhibition potency down to sub-micromolar. The identified molecules also inhibited the growth of different clinical presentations of C. trachomatis, one of the most common sexually transmitted disease worldwide.

2. Target-based screening against the P. aeruginosa virulence factor using enzymatic and biophysical assays. (Paper II-IV)

P. aeruginosa is a Gram-negative opportunistic pathogen with remarkable antibiotic resistance that is associated with a wide range of clinical infections. An alternative strategy to develop novel and selective antibacterials is to target the bacterial virulence factors, i.e. the ability of the bacteria to promote disease, thus ‘disarming’ the pathogens instead of killing them. P. aeruginosa employs its virulence factor, the type III secretion system (T3SS), to inject toxins (e.g. ExoS) into the eukaryotic cytosol. In one part of this thesis, we utilized enzymatic assay and identified inhibitors against the P. aeruginosa T3S toxin (ExoS). A follow up structure-activity relationship analysis was established and resulted in five (low micromolar) inhibitors of ExoS ADP-ribosylation enzymatic activity. In another part, we used surface plasmon resonance biophysical assay and identified small molecule binders of T3S translocation protein (PcrV). The primary SAR analysis was established and showed the antivirulence properties of these molecules and the potential to expand them further as novel antibacterials.

Place, publisher, year, edition, pages
Umeå: Umeå University, 2018. p. 95
Keywords
Antibacterials, antibiotics, small molecules, natural products, benzofuran, dihydrobenzofuran, the type III secretion system, Pseudomonas aeruginosa, Chlamydia trachomatis, phenotypic screening, high-throughput screening, surface plasmon resonance, drug discovery, bacterial toxins, enzyme inhibitors
National Category
Natural Sciences Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:umu:diva-150970 (URN)978-91-7601-917-7 (ISBN)
Public defence
2018-09-14, KB.E3.03 (Stora hörsalen), KBC-huset, Umeå, 09:00 (English)
Opponent
Supervisors
Funder
Swedish Foundation for Strategic Research , SSF, SB12-0022
Available from: 2018-08-24 Created: 2018-08-21 Last updated: 2018-08-21Bibliographically approved

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