High resolution 4D HPCH experiment for sequential assignment of (13)C-labeled RNAs via phosphodiester backbone.

The three-dimensional structure determination of RNAs by NMR spectroscopy requires sequential resonance assignment, often hampered by assignment ambiguities and limited dispersion of (1)H and (13)C chemical shifts, especially of C4'/H4'. Here we present a novel through-bond 4D HPCH NMR experiment involving phosphate backbone where C4'-H4' correlations are resolved along the (1)H3'-(31)P spectral planes. The experiment provides high peak resolution and effectively removes ambiguities encountered during assignments.

C4'/H4' selective, non-uniformly sampled 4D HC(P)CH experiment for sequential assignments of (13)C-labeled RNAs.

A through bond, C4'/H4' selective, "out and stay" type 4D HC(P)CH experiment is introduced which provides sequential connectivity via H4'(i)-C4'(i)-C4'(i-1)-H4'(i-1) correlations. The (31)P dimension (used in the conventional 3D HCP experiment) is replaced with evolution of better dispersed C4' dimension. The experiment fully utilizes (13)C-labeling of RNA by inclusion of two C4' evolution periods.

Structure determination of noncanonical RNA motifs guided by ¹H NMR chemical shifts.

Structured noncoding RNAs underlie fundamental cellular processes, but determining their three-dimensional structures remains challenging. We demonstrate that integrating ¹H NMR chemical shift data with Rosetta de novo modeling can be used to consistently determine high-resolution RNA structures. On a benchmark set of 23 noncanonical RNA motifs, including 11 'blind' targets, chemical-shift Rosetta for RNA (CS-Rosetta-RNA) recovered experimental structures with high accuracy (0.

NMR structure of 2'-O-(2-methoxyethyl) modified and C5-methylated RNA dodecamer duplex.

The solution-state structure of 2'-O-(2-methoxyethly) substituted dodecamer r(*CG*CGAA*U*U*CG*C)d(G), 2'-MOE RNA, with all cytosines and uracils methylated at the C5-position has been determined by NMR spectroscopy. The chemical modifications were used to improve the oligonucleotide's drug-like properties. The 2'-MOE group drives pseudorotational equilibrium of the ribofuranose moiety to the N-type conformation and supposedly results in structural preorganization leading to high affinity of a modified oligonucleotide towards its complementary biological target, improved pharmacokinetic and toxicological properties.

4D Non-uniformly sampled C,C-NOESY experiment for sequential assignment of 13C, 15N-labeled RNAs.

A 4D (13)C(aromatic),(13)C(ribose)-edited NOESY experiment is introduced to improve sequential assignment of non-coding RNA, often hampered by a limited dispersion of (1)H and (13)C chemical shifts. The (13)C-labeling of RNA is fully utilized by inclusion of two (13)C evolution periods. These dimensions provide enhanced dispersion of resonances in the 4D spectrum.

NMR studies of HAR1 RNA secondary structures reveal conformational dynamics in the human RNA.

Comparative genomics has shown that noncoding RNAs can display substantial differences between humans and chimpanzees. The human accelerated region 1 (HAR1) is a section in the human genome that exhibits the most strongly accelerated rate of nucleotide substitution in relation to the chimpanzee genome. It is associated with higher cognitive functions in human brains.

Solution structure of miRNA:mRNA complex.

The use of contemporary nuclear magnetic resonance (NMR) methods in the studies of model systems between microRNA (miRNA) and messenger RNA (mRNA) is reviewed. We describe our studies on structural features of 33-nt RNA model construct between let-7 miRNA and lin-41 mRNA at the second binding site. let-7 miRNA inhibits translation of lin-41 gene through formation of two complexes with the target sequence within 3' untranslated region of lin-41 mRNA in Caenorhabditis elegans.

NMR structure of the let-7 miRNA interacting with the site LCS1 of lin-41 mRNA from Caenorhabditis elegans.

We have determined the 3D structure of a 34-nt RNA construct, herein named LCS1co, which mimics the interaction of let-7 microRNA (miRNA) to one of its complementary binding sites, LCS1, in the 3'-untranslated region of lin-41 mRNA by solution-state NMR spectroscopy. let-7 miRNAs control the timing of development of the nematode Caenorhabditis elegans and are highly conserved in mammals. The sequence and structure of the two conserved let-7 complementary sites, LCS1 and LCS2, in the 3'-untranslated region of lin-41 mRNA are important for a proper downregulation of lin-41.

Solution structure of a let-7 miRNA:lin-41 mRNA complex from C. elegans.

Let-7 microRNA (miRNA) regulates heterochronic genes in developmental timing of the nematode Caenorhabditis elegans. Binding of miRNA to messenger RNA (mRNA) and structural features of the complex are crucial for gene silencing. We herein present the NMR solution structure of a model mimicking the interaction of let-7 miRNA with its complementary site (LCS 2) in the 3' untranslated region (3'-UTR) of the lin-41 mRNA.

Role of loop residues and cations on the formation and stability of dimeric DNA G-quadruplexes.

Formation of guanine-quadruplexes by four DNA oligonucleotides with common sequence dG4-loop-dG4 has been studied by a combination of NMR and UV spectroscopy. The loops consisted of 1',2'-dideoxyribose, propanediol, hexaethylene glycol, and thymine residues. The comparison of data on modified and parent oligonucleotides gave insight into the role of loop residues on formation and stability of dimeric G-quadruplexes.