RNA reactions one molecule at a time.
Much of the dynamics information is lost in bulk measurements because of the population averaging. Single-molecule methods measure one molecule at a time; they provide knowledge not obtainable by other means. In this article, we review the application of the two most widely used single-molecule methods--fluorescence resonance energy transfer (FRET) and force versus extension measurements--to several RNA reactions. First, we discuss folding/unfolding studies on a hairpin ribozyme that revealed multiple conformations of the RNA with distinct kinetics, and on a series of RNA pseudoknots, whose mechanical stabilities were found to show a strong correlation with their frameshifting efficiency during translation. We also discuss several RNA-related molecular motors. Single-molecule experiments revealed detailed mechanisms for the interaction of HIV reverse transcriptase and nucleic acid helicases (NS3 and RIG-1) with their substrates. Optical tweezers studies showed that translation of a single messenger RNA by a ribosome occurs by successive translocation-and-pause cycles. Single-molecule FRET experiments yielded important information on ribosome conformational changes and tRNA dynamics during translation. Overall, single-molecule experiments have been very valuable for understanding RNA reactions.
Tinoco
Ignacio
I
Department of Chemistry, University of California, Berkeley, California 94720-1460, USA. intinoco@lbl.gov
Chen
Gang
G
Qu
Xiaohui
X
D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D007700:Kinetics; D052898:Optical Tweezers; D012313:RNA / Q000737:chemistry*; D012270:Ribosomes / Q000737:chemistry*
D012313:RNA