RNA, one of essential components of life, is composed of four simple nucleotide building blocks that can assemble into complex forms. The basic nucleotides, namely U, A, G, C, can be decorated by various moieties that expand the structural and functional capacities. RNA modifications are frequently present in many different types of RNA. In our work we combine CryoEM, biophysical, and biochemical tools with the aim to structurally characterize the basis of selected RNA modifications as well as the protein complexes responsible for the modifications.
The intent of this presentation is to introduce our biophysical platform consisting of the Prometheus Panta and Monolith series of instruments from NanoTemper Technologies. The presentation will cover the biophysical background of the Monolith, used routinely in our laboratory to study interactions of the protein-RNA complexes. In addition, I will introduce the Prometheus Panta technology and its application to characterize protein properties. This technology allows us to characterize colloidal and conformational protein properties; assessing not only the quality of our protein purification pipeline but also effects of clinical point mutations on the protein conformational stability. The presentation will include an example of our structural and biophysical characterization of the protein-RNA complex involved in writing a commonly occurring RNA modification.