Single-Molecule Fluorescence Energy Transfer Assays for the Characterization of Reaction Pathways of miRNA-Argonaute Complex

Argonaute proteins are key components of the microRNA-induced silencing complexes (miRISCs) that mediate the posttranscriptional gene silencing of microRNAs and small interfering RNA (siRNAs). The complex reaction mechanism of miRISC is expected to be characterized by tracing the reaction pathways of miRISC at the single-molecule level in real time. In this chapter, we describe single-molecule fluorescence resonance energy transfer (FRET) assays to observe the target binding and reaction pathways of miRISC composed of a recombinant Argonaute and a small RNA.