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Updated: Mar 22, 2026

Fluorescence Assays for the Study of Mycobacterium tuberculosis Interaction with the Immune Receptor SLAMF1
Published on: February 28, 2025
FRET reveals multiple interaction states between two component signalling system proteins of M. tuberculosis
Ruchi Agrawal1, Prem Kumar V1, Harini Ramanan1
1Department of Molecular Reproduction, Development and Genetics, Indian Institute of Science, Bangalore, India.
Background:
Two component signalling involves interaction between sensor kinase (SK) and response regulator (RR) proteins which depends on their phosphorylation status.
Methods:
In this study we report the development of an in vitro FRET assay for studying interaction between fluorescently tagged SK and RR proteins.
Results:
Using TCS proteins of Mycobacterium tuberculosis, we demonstrate that phosphorylation status of SK affects the SK-RR interaction, which varies from one TCS to another. The observation was strengthened by recordings from mutant SK and RR proteins. The assay retained the specificity/crosstalk potential of the participating proteins and reflected the inherent phosphotransfer potentials.
Conclusions:
SK and RR proteins interact with each other in unphosphorylated state and the phosphorylation affects the interaction between SK and RR, which was reflected as reduction in FRET ratio.
General Significance:
A non-radioactive, in vitro FRET based assay is reported, which can be utilized for studying genome-wide partner screening, identifying crosstalk or specificity in TCSs.
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