Fluorescence spectroscopy is a sensitive technique for detecting protein-protein, protein-RNA, and RNA-RNA interactions, requiring only nanomolar concentrations of labeled components. Fluorescence anisotropy provides information about the assembly of multi-subunit proteins, while molecular beacons provide a sensitive and quantitative reporter for base pairing between complementary RNAs. Here we present a detailed protocol for labeling Hfq protein with cyanine 3-maleimide and dansyl chloride to study the protein oligomerization and RNA binding by semi-native polyacrylamide gel electrophoresis (PAGE) and fluorescence anisotropy. We also present a detailed protocol for measuring the rate of annealing between a molecular beacon and a target RNA in the presence of Hfq using a stopped-flow spectrometer.
CITATION STYLE
Panja, S., & Woodson, S. A. (2015). Fluorescence reporters for Hfq oligomerization and RNA annealing. In RNA Remodeling Proteins: Methods and Protocols (pp. 369–383). Springer New York. https://doi.org/10.1007/978-1-4939-2214-7_22
Mendeley helps you to discover research relevant for your work.