Experimental and computational investigations of RNA duplexes containing N7-regioisomers of adenosine and LNA-adenosine.

Although glycosidic bonds in purines typically involve the N9 position, the chemical synthesis of adenosine produces N7-ribofuranosyladenine (7A) as a kinetically favorable ribosylation product. Similarly, in the synthesis of LNA-adenosine (AL), a minor product, N7-LNA-adenosine (7AL), is observed. While extensive research has focused on investigating the properties of N9-regioisomers of adenosine, 7A has been largely overlooked and considered as a side-product.

The oligonucleotides containing N7-regioisomer of guanosine: influence on thermodynamic properties and structure of RNA duplexes.

During the chemical synthesis of the purine riboside, N7-regioisomer is kinetically formed, whereas N9-regioisomer is a thermodynamically formed product. We have studied the effect of substituting N9-regioisomer of guanosine with its N7-regioisomer (N7-guanosine, 7G) at a central position of several RNA duplexes. We found that this single substitution by 7G severely diminished their thermodynamic stabilities when 7G paired with C and U, but remarkably, led to a significant amount of stabilization in most of the duplexes when forming mismatches with G and A.

The 8-17 DNAzyme can operate in a single active structure regardless of metal ion cofactor.

DNAzymes - synthetic enzymes made of DNA - have long attracted attention as RNA-targeting therapeutic agents. Yet, as of now, no DNAzyme-based drug has been approved, partially due to our lacking understanding of their molecular mode of action. In this work we report the solution structure of 8-17 DNAzyme bound to a Zn ion solved through NMR spectroscopy.

Comparison of the Backbone Dynamics of Dehaloperoxidase-Hemoglobin Isoenzymes.

Dehaloperoxidase (DHP) is a multifunctional hemeprotein with a functional switch generally regulated by the chemical class of the substrate. Its two isoforms, DHP-A and DHP-B, differ by only five amino acids and have an almost identical protein fold. However, the catalytic efficiency of DHP-B for oxidation by a peroxidase mechanism ranges from 2- to 6-fold greater than that of DHP-A depending on the conditions.

Solution Structure of a Lanthanide-binding DNA Aptamer Determined Using High Quality pseudocontact shift restraints.

In this contribution we report the high-resolution NMR structure of a recently identified lanthanide-binding aptamer (LnA). We demonstrate that the rigid lanthanide binding by LnA allows for the measurement of anisotropic paramagnetic NMR restraints which to date remain largely inaccessible for nucleic acids. One type of such restraints - pseudocontact shifts (PCS) induced by four different paramagnetic lanthanides - was extensively used throughout the current structure determination study and the measured PCS turned out to be exceptionally well reproduced by the final aptamer structure.

Impact of a Single Nucleotide Change or Non-Nucleoside Modifications in G-Rich Region on the Quadruplex-Duplex Hybrid Formation.

In this paper, a method to discriminate between two target RNA sequences that differ by one nucleotide only is presented. The method relies on the formation of alternative structures, i.e.

RNA and DNA G-quadruplexes bind to human dicer and inhibit its activity.

Guanine (G)-rich single-stranded nucleic acids can adopt G-quadruplex structures. Accumulating evidence indicates that G-quadruplexes serve important regulatory roles in fundamental biological processes such as DNA replication, transcription, and translation, while aberrant G-quadruplex formation is linked to genome instability and cancer. Understanding the biological functions played by G-quadruplexes requires detailed knowledge of their protein interactome.

Formation of an RNA Quadruplex-Duplex Hybrid in Living Cells between mRNA of the Epidermal Growth Factor Receptor (EGFR) and a G-Rich Antisense Oligoribonucleotide.

Antisense DNA oligonucleotides, short interfering RNAs (siRNAs), and CRISPR/Cas9 genetic tools are the most useful therapeutic nucleic acids regulating gene expression based on the antisense specificity towards messenger RNA. Here, we present an effective novel strategy for inhibiting translation based on the antisense-controlled formation of an RNA quadruplex-duplex hybrid (QDH) between a G-rich RNA antisense oligoribonucleotide (Q-ASO) and specific mRNA, comprising two distant G-tracts. We selected epidermal growth factor receptor (EGFR) as a well-established target protein in anticancer therapy.

The origin of the high stability of 3'-terminal uridine tetrads: contributions of hydrogen bonding, stacking interactions, and steric factors evaluated using modified oligonucleotide analogs.

RNA G-quadruplexes fold almost exclusively into parallel-stranded structures and thus display much less structural diversity than their DNA counterparts. However, also among RNA G-quadruplexes peculiar structural elements can be found which are capable of reshaping the physico-chemical properties of the folded structure. A striking example is provided by a uridine tetrad (U-tetrad) placed on the 3'-terminus of the tetramolecular G-quadruplex.

A strong preference for the TA/TA dinucleotide step discovered for an acridine-based, potent antitumor dsDNA intercalator, C-1305: NMR-driven structural and sequence-specificity studies.

Triazoloacridinone C-1305, a potent antitumor agent recommended for Phase I clinical trials, exhibits high activity towards a wide range of experimental colon carcinomas, in many cases associated with complete tumor regression. C-1305 is a well-established dsDNA intercalator, yet no information on its mode of binding into DNA is available to date. Herein, we present the NMR-driven and MD-refined reconstruction of the 3D structures of the d(CGATATCG):C-1305 and d(CCCTAGGG):C-1305 non-covalent adducts.

Computational and NMR studies of RNA duplexes with an internal pseudouridine-adenosine base pair.

Pseudouridine (Ψ) is the most common chemical modification present in RNA. In general, Ψ increases the thermodynamic stability of RNA. However, the degree of stabilization depends on the sequence and structural context.

Thermodynamic and structural contributions of the 6-thioguanosine residue to helical properties of RNA.

Thionucleotides, especially 4-thiouridine and 6-thioguanosine, are photosensitive molecules that photocrosslink to both proteins and nucleic acids, and this feature is a major reason for their application in various investigations. To get insight into the thermodynamic and structural contributions of 6-thioguanosine to the properties of RNA duplexes a systematic study was performed. In a series of RNA duplexes, selected guanosine residues located in G-C base pairs, mismatches (G-G, G-U, and G-A), or 5' and 3'-dangling ends were replaced with 6-thioguanosine.

Unraveling the structural basis for the exceptional stability of RNA G-quadruplexes capped by a uridine tetrad at the 3' terminus.

Uridine tetrads (U-tetrads) are a structural element encountered in RNA G-quadruplexes, for example, in the structures formed by the biologically relevant human telomeric repeat RNA. For these molecules, an unexpectedly strong stabilizing influence of a U-tetrad forming at the 3' terminus of a quadruplex was reported. Here we present the high-resolution solution NMR structure of the r(UGGUGGU) quadruplex which, in our opinion, provides an explanation for this stabilization.

Metal-cation regulation of enzyme dynamics is a key factor influencing the activity of S-adenosyl-L-homocysteine hydrolase from Pseudomonas aeruginosa.

S-adenosyl-L-homocysteine hydrolase from Pseudomonas aeruginosa (PaSAHase) coordinates one K ion and one Zn ion in the substrate binding area. The cations affect the enzymatic activity and substrate binding but the molecular mechanisms of their action are unknown. Enzymatic and isothermal titration calorimetry studies demonstrated that the K ions stimulate the highest activity and strongest ligand binding in comparison to other alkali cations, while the Zn ions inhibit the enzyme activity.

Dynamics of dehaloperoxidase-hemoglobin A derived from NMR relaxation spectroscopy and molecular dynamics simulation.

Dehaloperoxidase-hemoglobin is the first hemoglobin identified with biologically-relevant oxidative functions, which include peroxidase, peroxygenase and oxidase activities. Herein we report a study of the protein backbone dynamics of DHP using heteronuclear NMR relaxation methods and molecular dynamics (MD) simulations to address the role of protein dynamics in switching from one function to another. The results show that DHP's backbone helical regions and turns have average order parameters of S = 0.

Author Correction: Identification of functional tetramolecular RNA G-quadruplexes derived from transfer RNAs.

The original version of this Article contained an error in the spelling of the author Steven M. Coyne, which was incorrectly given as Stephen M. Coyne.

Identification of functional tetramolecular RNA G-quadruplexes derived from transfer RNAs.

RNA G-quadruplex (RG4) structures are involved in multiple biological processes. Recent genome-wide analyses of human mRNA transcriptome identified thousands of putative intramolecular RG4s that readily assemble in vitro but shown to be unfolded in vivo. Previously, we have shown that mature cytoplasmic tRNAs are cleaved during stress response to produce tRNA fragments that function to repress translation in vivo.

Effects of G-quadruplex topology on translational inhibition by tRNA fragments in mammalian and plant systems in vitro.

The folding of tRNA fragments (tRFs) into G-quadruplex structures and the implications of G-quadruplexes in translational inhibition have been studied mainly in mammalian systems. To increase our knowledge of these phenomena, we determined the influence of human and plant tRFs and model G-quadruplexes on translation in rabbit reticulocyte lysate and wheat germ extract. The efficiency of translational inhibition in the mammalian system was strongly associated with the type of G-quadruplex topology.

A hydantoin isoform of cyclic N6-threonylcarbamoyladenosine (ct6A) is present in tRNAs.

N6-Threonylcarbamoyladenosine (t6A) and its derivatives are universally conserved modified nucleosides found at position 37, 3΄ adjacent to the anticodon in tRNAs responsible for ANN codons. These modifications have pleiotropic functions of tRNAs in decoding and protein synthesis. In certain species of bacteria, fungi, plants and protists, t6A is further modified to the cyclic t6A (ct6A) via dehydration catalyzed by TcdA.

Overview of the RNA G-quadruplex structures.

G-quadruplexes are non-canonical secondary structures which may be formed by guanine rich sequences, both in vitro and in living cells. The number of biological functions assigned to these structural motifs has grown rapidly since the discovery of their involvement in the telomere maintenance. Knowledge of the G-quadruplexes' three-dimensional structures plays an important role in understanding of their conformational diversity, physiological functions, and in the design of novel drugs targeting the G-quadruplexes.

Thermodynamic Features of Structural Motifs Formed by β-L-RNA.

This is the first report to provide comprehensive thermodynamic and structural data concerning duplex, hairpin, quadruplex and i-motif structures in β-L-RNA series. Herein we confirm that, within the limits of experimental error, the thermodynamic stability of enantiomeric structural motifs is the same as that of naturally occurring D-RNA counterparts. In addition, formation of D-RNA/L-RNA heterochiral duplexes is also observed; however, their thermodynamic stability is significantly reduced in reference to homochiral D-RNA duplexes.

Structural characterization of a dimer of RNA duplexes composed of 8-bromoguanosine modified CGG trinucleotide repeats: a novel architecture of RNA quadruplexes.

Fragile X syndrome and fragile X-associated tremor/ataxia syndrome (FXTAS) are neurodegenerative disorders caused by the pathogenic expansion of CGG triplet repeats in the FMR1 gene. FXTAS is likely to be caused by a 'toxic' gain-of-function of the FMR1 mRNA. We provide evidence for the existence of a novel quadruplex architecture comprising CGG repeats.

Photochemical studies and nanomolar photodynamic activities of phthalocyanines functionalized with 1,4,7-trioxanonyl moieties at their non-peripheral positions.

Manganese(III), cobalt(II), copper(II), magnesium(II), zinc(II) and metal-free phthalocyanines, possessing 1,4,7-trioxanonyl substituents, at their non-peripheral positions, were subjected to photochemical, photodynamic and biological activity studies. Demetallated phthalocyanine and its metallated d-block analogues, with copper(II), cobalt(II), manganese(III) chloride, were found to be less efficient singlet oxygen generators in comparison to the zinc(II) analogue and zinc(II) phthalocyanine reference. Irradiation of several phthalocyanines for short time periods resulted in a substantially increased cytostatic activity against both suspension (leukemic/lymphoma at 85nM) and solid (cervix carcinoma at 72nM and melanoma at 81nM) tumour cell lines (up to 200-fold).

Fusion of the H NMR data of serum, urine and exhaled breath condensate in order to discriminate chronic obstructive pulmonary disease and obstructive sleep apnea syndrome.

Chronic obstructive pulmonary disease, COPD, affects the condition of the entire human organism and causes multiple comorbidities. Pathological lung changes lead to quantitative changes in the composition of the metabolites in different body fluids. The obstructive sleep apnea syndrome, OSAS, occurs in conjunction with chronic obstructive pulmonary disease in about 10-20 % of individuals who have COPD.