Chemo-enzymatic synthesis of site-specific isotopically labeled nucleotides for use in NMR resonance assignment, dynamics and structural characterizations

Longhini, Andrew P.; LeBlanc, Regan M.; Becette, Owen; Salguero, Carolina; Wunderlich, Christoph H.; Johnson, Bruce A.; D'Souza, Victoria M.; Kreutz, Christoph; Dayie, T. Kwaku

A price to pay for not using spectroscopic tools to minimize the 13 C-13 C coupling problem is that the number of probe sites is now limited to the labeled sites. Nonetheless, it still remains useful because our method allows for very rapid accumulation of chemical shifts, a set of parameters that are easily and accurately measured and available at very early stages of NMR data analyses. Thus by measuring various chemical shifts (H1 /C1 , H2 /C2 , H5 ,H5 /C5 , H2/C2, C4, H5/C5, H6/C6, H8/C8, N1, N3, N7, N9), we think the availability of such parameters will facilitate chemical shift based structure calculations of RNA, especially for constructing structural models for transiently and sparsely populated RNA states as has been done, so far, only for proteins (97) .