Combining two optimized and affordable methods to assign chemoreceptors to a specific signal - Archive ouverte HAL Access content directly
Journal Articles Analytical Biochemistry Year : 2021

Combining two optimized and affordable methods to assign chemoreceptors to a specific signal

Abstract

Chemotaxis allows bacteria to detect specific compounds and move accordingly. This pathway involves signal detection by chemoreceptors (MCPs). Attributing a chemoreceptor to a ligand is difficult because there is a lot of redundancy in the MCPs that recognize a single ligand. We propose a methodology to define which chemoreceptors bind a given ligand. First, an MCP is overproduced to increase sensitivity to the ligand(s) it recognizes, thus promoting accumulation of cells around an agarose plug containing a low attractant concentration. Second, the ligand-binding domain (LBD) of the chemoreceptor is fused to maltose-binding protein (MBP), which facilitates purification and provides a control for a thermal shift assay (TSA). An increase in the melting temperature of the LBD in the presence of the ligand indicates that the chemoreceptor directly binds it. We showed that overexpression of two Shewanella oneidensis chemoreceptors (SO_0987 and SO_1056) promoted swimming toward an agarose plug containing a low concentration of chromate. The LBD of each of the two chemoreceptors was fused to MBP. A TSA revealed that only the LBD from SO_1056 had its melting temperature increased by chromate. In conclusion, we describe an efficient approach to define chemoreceptor-ligand pairs before undertaking more-sophisticated biochemical and structural studies.
Fichier principal
Vignette du fichier
S0003269721000403.pdf (1.36 Mo) Télécharger le fichier
Origin Files produced by the author(s)

Dates and versions

hal-03195889 , version 1 (10-03-2023)

Licence

Identifiers

Cite

Anne Boyeldieu, Amine Ali Chaouche, Vincent Méjean, Cécile Jourlin-Castelli. Combining two optimized and affordable methods to assign chemoreceptors to a specific signal. Analytical Biochemistry, 2021, 620, pp.114139. ⟨10.1016/j.ab.2021.114139⟩. ⟨hal-03195889⟩

Collections

CNRS UNIV-AMU
43 View
23 Download

Altmetric

Share

Gmail Mastodon Facebook X LinkedIn More