RNA synthesis and purification for structural studies

Abstract

RNAs play pivotal roles in the cell, ranging from catalysis (e.g., RNase P), acting as adaptor molecule (tRNA) to regulation (e.g., riboswitches). Precise understanding of its three-dimensional structures has given unprecedented insight into the molecular basis for all of these processes. Nevertheless, structural studies on RNA are still limited by the very special nature of this polymer. The most common methods for the determination of 3D RNA structures are NMR and X-ray crystallography. Both methods have their own set of requirements and give different amounts of information about the target RNA. For structural studies, the major bottleneck is usually obtaining large amounts of highly pure and homogeneously folded RNA. Especially for X-ray crystallography it can be necessary to screen a large number of variants to obtain well-ordered single crystals. In this mini-review we give an overview about strategies for the design, in vitro production, and purification of RNA for structural studies.

Keywords: :

• in vitro transcription
• structural biology
• X-ray
• NMR
• ribozymes

10.4161/rna.28076

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.

Acknowledgments

We thank Achim Dickmanns and Thomas Monecke for critical reading of the manuscript, and the DFG SPP1258 for funding.