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Light-activated chemical probing of nucleobase solvent accessibility inside cells.

Abstract

The discovery of functional RNAs that are critical for normal and disease physiology continues to expand at a breakneck pace. Many RNA functions are controlled by the formation of specific structures, and an understanding of each structural component is necessary to elucidate its function. Measuring solvent accessibility intracellularly with experimental ease is an unmet need in the field. Here, we present a novel method for probing nucleobase solvent accessibility, Light Activated Structural Examination of RNA (LASER). LASER depends on light activation of a small molecule, nicotinoyl azide (NAz), to measure solvent accessibility of purine nucleobases. In vitro, this technique accurately monitors solvent accessibility and identifies rapid structural changes resulting from ligand binding in a metabolite-responsive RNA. LASER probing can further identify cellular RNA-protein interactions and unique intracellular RNA structures. Our photoactivation technique provides an adaptable framework to structurally characterize solvent accessibility of RNA in many environments.

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