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Chemiluminescent sensors for quantitation of the bacterial second messenger cyclic di-GMP.

Authors :
Anderson WA
Dippel AB
Maiden MM
Waters CM
Hammond MC
Source :
Methods in enzymology [Methods Enzymol] 2020; Vol. 640, pp. 83-104. Date of Electronic Publication: 2020 May 30.
Publication Year :
2020

Abstract

Chemiluminescent biosensors have been developed and broadly applied to mammalian cell systems for studying intracellular signaling networks. For bacteria, biosensors have largely relied on fluorescence-based systems for quantitating signaling molecules, but these designs can encounter issues in complex environments due to their reliance on external illumination. In order to circumvent these issues, we designed the first ratiometric chemiluminescent biosensors for studying a key bacterial second messenger, cyclic di-GMP. We have shown recently that these biosensors function both in vitro and in vivo for detecting changes in cyclic di-GMP levels. In this chapter, we present a practical and broadly applicable method for high-throughput quantitation of cyclic di-GMP in bacterial cell extracts using the high affinity biosensor tVYN-TmΔ that could serve as the "Bradford assay" equivalent for this bacterial signaling molecule.<br /> (© 2020 Elsevier Inc. All rights reserved.)

Details

Language :
English
ISSN :
1557-7988
Volume :
640
Database :
MEDLINE
Journal :
Methods in enzymology
Publication Type :
Academic Journal
Accession number :
32560807
Full Text :
https://doi.org/10.1016/bs.mie.2020.04.004