Microscale Thermophoresis to Study RNA-RNA Binding Affinity

Abstract

Evaluation of RNA-RNA binding is crucial for in vitro studying of molecular mechanisms, for example, the interaction of noncoding RNAs (ncRNAs) with their respective targets. In recent years, the method of microscale thermophoresis (MST) has been developed, which is based on the physical phenomenon of thermophoresis (Ludwig-Soret Effect), defined as the migration of a molecule in a solution in response to a macroscopic temperature gradient. The method enables the fast detection and characterization of biophysical interaction between molecules, with the fundamental advantage that only small amounts of target and ligand are required. Here, we describe the characterization of RNA-RNA binding affinity using the example of the sRNA₄₁ from Methanosarcina mazei and its native target, the 5' UTR of mRNA-MM2089, the first gene of the operon encoding the acetyl-CoA decarboxylase/synthase complex.

Keywords: MST; Microscale thermophoresis; RNA–RNA binding affinity; sRNA.