Format

Send to

Choose Destination
J Am Chem Soc. 2019 Jan 30;141(4):1430-1434. doi: 10.1021/jacs.8b11384. Epub 2019 Jan 22.

Long-Range RNA Structural Information via a Paramagnetically Tagged Reporter Protein.

Author information

1
Laboratory of Structural Biophysics, Biochemistry and Biophysics Center, National Heart, Lung and Blood Institute , National Institutes of Health , Bethesda , Maryland 20892 , United States.
2
Office of Intramural Research, Center for Information Technology, National Institutes of Health , Bethesda , Maryland 20892 , United States.

Abstract

NMR has provided a wealth of structural and dynamical information for RNA molecules of up to ∼50 nucleotides, but its application to larger RNAs has been hampered in part by difficulties establishing global structural features. A potential solution involves measurement of NMR perturbations after site-specific paramagnetic labeling. Although the approach works well for proteins, the inability to place the label at specific sites has prevented its application to larger RNAs transcribed in vitro. Here, we present a strategy in which RNA loop residues are modified to promote binding to a paramagnetically tagged reporter protein. Lanthanide-induced pseudocontact shifts are demonstrated for a 232-nucleotide RNA bound to tagged derivatives of the spliceosomal U1A RNA-binding domain. Further, the method is validated with a 36-nucleotide RNA for which measured NMR values agreed with predictions based on the previously known protein and RNA structures. The ability to readily insert U1A binding sites into ubiquitous hairpin and/or loop structures should make this approach broadly applicable for the atomic-level study of large RNAs.

PMID:
30652860
PMCID:
PMC6572783
[Available on 2020-01-30]
DOI:
10.1021/jacs.8b11384

Supplemental Content

Full text links

Icon for American Chemical Society
Loading ...
Support Center