Self‐Assembly of DNA and RNA Building Blocks Explored by Nitrogen‐14 NMR Crystallography: Structure and Dynamics

Abstract

The isotopic enrichment of nucleic acids with nitrogen-15 is often carried out by solid-phase synthesis of oligonucleotides using phosphoramidite precursors that are synthetically demanding and expensive. These synthetic challenges, combined with the overlap of chemical shifts, explain the lag of nitrogen-15 NMR studies of nucleic acids behind those of proteins. For the structural characterization of DNA and RNA-related systems, new NMR methods that exploit the naturally occurring 99.9 % abundant nitrogen-14 isotope are therefore highly desirable. In this study, we have investigated nitrogen-14 spectra of selfassembled quartets based on the nucleobase guanine in the solid state by means of magic-angle spinning NMR spectro-scopy. The network of dipolar proton-nitrogen couplings between neighboring stacked purine units is probed by 2D spectra based on 1 H! 14 N! 1 H double cross-polarization. Interplane dipolar contacts are identified between the stacked G quartets. The assignment is supported by density functional theory (DFT) calculations of the anisotropic chemical shifts and quadrupolar parameters. The experimental spectra are fully consistent with internuclear distances obtained in silico. Averaging of chemical shifts due to internal motions can be interpreted by semiempirical calculations. This method can easily be extended to synthetic G quartets based on nucleobase or nucleoside analogs and potentially to oligonucleotides.[a] Dr.

In-Cell Structural Biology by NMR: The Benefits of the Atomic Scale

“…At first glance, this could ruin the enthusiasm about in-cell NMR: the most abundant isotopes of carbon ( 12 C, natural abundance 98.9%), oxygen ( 16 O, 99.8%) or sulfur ( 32 S, 95%) have a null spin. Moreover, 14 N (99.6% of nitrogen) has unfavorable spin quantum number and quadrupolar moment values, which makes it poorly detectable with the current NMR techniques (Except for the small, symmetric molecules NH 4 + and NO 3 – or using ssNMR approaches still far from routine and from relevant cellular concentration ranges). , In-cell NMR spectroscopists turned this drawback into a blessing: By delivering in cells molecules enriched in 13 C and 15 N, two isotopes with NMR friendly characteristics, they can execute 13 C- or 15 N-editing NMR techniques in an almost blank cellular background (Figure ). In this regard, non-natural 19 F-containing amino acids or nucleic acids can also be convenient, because cells do not contain any fluorine.…”

“…Moreover, 14 N (99.6% of nitrogen) has unfavorable spin quantum number and quadrupolar moment values, which makes it poorly detectable with the current NMR techniques (Except for the small, symmetric molecules NH 4 + and NO 3 − or using ssNMR approaches still far from routine and from relevant cellular concentration ranges). 80,81 In-cell NMR spectroscopists turned this drawback into a blessing: By delivering in cells molecules enriched in 13 C and 15 N, two isotopes with NMR friendly characteristics, they can execute 13 C-or 15 N-editing NMR techniques in an almost blank cellular background (Figure 3). In this regard, non-natural 19 F-containing amino acids or nucleic acids can also be convenient, because cells do not contain any fluorine.…”

Enzymatic construction of metal-mediated nucleic acid base pairs

“…28 The use of 14 N NMR to investigate the interaction of metal cations with nucleic acids, particularly in the context of metal base pairs, would be a highly interesting and versatile tool but is not sufficiently developed as yet . 101 In this context, Johannsen et al published in 2010 the NMR solution structure of a self-complementary DNA oligonucleotide containing three consecutive imidazole nucleotides in its centre. The structure has been reported both in the absence and presence of Ag I ions.…”

Combining heteronuclear correlation NMR with spin-diffusion to detect relayed Cl–H–H and N–H–H proximities in molecular solids

“…For 2D experiments involving both spin-half and quadrupolar nuclei, heteronuclear multiple-quantum coherence (HMQC)-type or insensitive nuclei enhancement by polarization transfer (INEPT)-type experiments have been used to efficiently transfer the polarization from spin-half nuclei to quadrupolar nuclei [19][20][21][22][23][24][25]. For example, N-H proximities can be probed at natural abundance ( 14 N spin 1, 99.6% natural isotopic abundance) using J-couplings (J-HMQC), dipolar couplings (D-HMQC), and using overtone (OT) 14 N transitions [23,[25][26][27][28][29][30][31][32][33]. Specifically, 2D HMQC experiments have been applied to characterize 14 N- 1 H proximities [28,[34][35][36][37][38] as well as 35 Cl-1 H proximities [38][39][40][41][42] in small molecules and pharmaceutical solids.…”