Establishment of an In Bacterio Assay for the Assessment of Carbon Storage Regulator A (CsrA) Inhibitors.
anti-infectives
carbon storage regulator A
reporter gene assay
repressor of secondary metabolites A
virulence modulation
Journal
Chembiochem : a European journal of chemical biology
ISSN: 1439-7633
Titre abrégé: Chembiochem
Pays: Germany
ID NLM: 100937360
Informations de publication
Date de publication:
15 08 2023
15 08 2023
Historique:
revised:
12
07
2023
received:
17
05
2023
medline:
17
8
2023
pubmed:
12
7
2023
entrez:
12
7
2023
Statut:
ppublish
Résumé
Polymicrobial infections involving various combinations of microorganisms, such as Escherichia, Pseudomonas, or Yersinia, can lead to acute and chronic diseases in for example the gastrointestinal and respiratory tracts. Our aim is to modulate microbial communities by targeting the posttranscriptional regulator system called carbon storage regulator A (CsrA) (or also repressor of secondary metabolites (RsmA)). In previous studies, we identified easily accessible CsrA binding scaffolds and macrocyclic CsrA binding peptides through biophysical screening and phage display technology. However, due to the lack of an appropriate in bacterio assay to evaluate the cellular effects of these inhibitor hits, the focus of the present study is to establish an in bacterio assay capable of probing and quantifying the impact on CsrA-regulated cellular mechanisms. We have successfully developed an assay based on a luciferase reporter gene assay, which in combination with a qPCR expression gene assay, allows for the monitoring of expression levels of different downstream targets of CsrA. The chaperone protein CesT was used as a suitable positive control for the assay, and in time-dependent experiments, we observed a CesT-mediated increase in bioluminescence over time. By this means, the cellular on-target effects of non-bactericidal/non-bacteriostatic virulence modulating compounds targeting CsrA/RsmA can be evaluated.
Identifiants
pubmed: 37435861
doi: 10.1002/cbic.202300369
doi:
Substances chimiques
Escherichia coli Proteins
0
Carbon
7440-44-0
RNA-Binding Proteins
0
Bacterial Proteins
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
e202300369Informations de copyright
© 2023 The Authors. ChemBioChem published by Wiley-VCH GmbH.
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