Use of ¹³C as an Indirect Tag in ¹⁵N Specifically Labeled Nucleosides. Syntheses of [8-¹³C-1,7,NH₂-¹⁵N₃]Adenosine, -Guanosine, and Their Deoxy Analogues

Abstract

We have previously reported the use of a 13C tag at the C2 of 15N-multilabeled purine nucleosides to distinguish the adjacent-labeled 15N atoms from those in an untagged nucleoside. We now introduce the use of an indirect tag at the C8 of 15N7-labeled purine nucleosides. This tag allows unambiguous differentiation between a pair of 15N7-labeled purines in which only one is 13C8 labeled. Although the very small C8-N7 coupling (<1 Hz) precludes its direct detection in 1D 15N spectra, 2D 1H-15N NMR experiments display the large C8-H8 coupling (>200 Hz) because H8 is coupled to both N7 and C8. The 13C8 atom is introduced by means of a ring closure of the exocyclic amino groups of a pyrimidinone using [13C]sodium ethyl xanthate. Here, we present methods for the syntheses of [8-13C-1,7,NH2-15N3]adenosine, -guanosine, and their deoxy analogues.

Mapping Structurally Defined Guanine Oxidation Products along DNA Duplexes: Influence of Local Sequence Context and Endogenous Cytosine Methylation

“…15 N 3 , 13 C 1 -dG and 15 N 3 , 13 C 1 -dG phosphoramidites were prepared as described previously (dG: 2′-deoxyguanosine). 55 …”

“…15 N 3 , 13 C 1 -dG was introduced at specified positions by solid phase synthesis using 1,7,NH 2 - 15 N-2- 13 C-dG phosphoramidite. 55,56 DNA strands containing phenylpyrrolo-C were prepared as described previously. 57,58 All DNA oligomers were purified by HPLC to 99+% purity (Figure S2 in the Supporting Information) and characterized by UV spectroscopy and MALDI mass spectrometry as described previously.…”

Post-synthetic and site-specific modification of endocyclic nitrogen atoms of purines in DNA and its potential for biological and structural studies

“…Expensive isotopes, such as 15 N, can be cost-effectively introduced onto the specified base by treatment with appropriate reagents in the final step (13,14). Such site-specific 15 N labelling at the oligomer level can be viewed as a complementary approach with those used for initial synthesis of 15 N-labelled nucleosides and subsequent preparation of labelled DNA (15,16). …”

Heavy atom labeled nucleotides for measurement of kinetic isotope effects

“…It should be noted that we presented this collection of nucleic acid analogs from a perspective of RNA catalysis. Consequently, we excluded methodologies for heavy atom nucleosides that have no obvious application for RNA catalysis related isotope effect studies including [7- 15 N] purines [51]. However, there exist many nucleic acid modifying enzymes that play essential roles in biology for which mechanistic enzymology has so far provided only limited understanding.…”