Chemo-enzymatic labeling for rapid assignment of RNA molecules

• Published in: Methods (San Diego, Calif.) Calif.), 2016-07, Vol.103, p.11-17
• Main Authors: Longhini, Andrew P., LeBlanc, Regan M., Dayie, T. Kwaku
• Format: Article
• Language: English
• Subjects: A-site; Amides - chemistry; Atomic-, site-selective isotopic labels; Base Sequence; Filtered/edited NOESY; Inverted Repeat Sequences; Isotope Labeling; Magnetic Resonance Spectroscopy; NMR; Phosphoric Acids - chemistry; Purines - chemistry; Pyrimidines - chemistry; RNA; RNA - chemical synthesis
• ISSN: 1046-2023; 1095-9130
• DOI: 10.1016/j.ymeth.2016.03.015

[Display omitted] •Tools exist to tackle problems of NMR chemical shift overlap and rapid signal loss.•Chemoenzymatic synthesis affords flexible, cost-effective means to make labeled NTPs.•Atom-specific isotopic enrichment enable rapid NMR assignment of RNA helical regions.•Hybrid phosphoramidite-selective labeling will be effective for solving 3D structure.•Labeled nucleotides amenable to segmental, phosphoramidite, and PLOR technologies. Even though Nuclear Magnetic Resonance (NMR) spectroscopy is one of the few techniques capable of determining atomic resolution structures of RNA, it is constrained by two major problems of chemical shift overlap of resonances and rapid signal loss due to line broadening. Emerging tools to tackle these problems include synthesis of atom specifically labeled or chemically modified nucleotides. Herein we review the synthesis of these nucleotides, the design and production of appropriate RNA samples, and the application and analysis of the NMR experiments that take advantage of these labels.