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Phytosterols are known to reduce plasma cholesterol levels and thereby reduce cardiovascular risk. Studies conducted on human and animal models have demonstrated that these compounds have also anti-inflammatory effects. Recently, an experimental colitis model (dextran sulphate sodium-induced) has shown that pre-treatment with phytosterols decreases infiltration of inflammatory cells and accelerates mucosal healing. This study aims to understand the mechanism underlying the colitis by analysing the end-products of the metabolism in distal colon and liver excised from the same mice used in the previous work. In particular, an unsupervised gas chromatography-mass spectrometry (GC-MS) and NMR based metabolomics approach was employed to identify the metabolic pathways perturbed by the dextran sodium sulphate (DSS) insult (i.e. Krebs cycle, carbohydrate, amino acids, and nucleotide metabolism). Interestingly, phytosterols were able to restore the homeostatic equilibrium of the hepatic and colonic metabolome. [ABSTRACT FROM AUTHOR]

The DNA sequence d(GGGGTTTTGGGG) consists of 1.5 units of the repeat in telomeres of Oxytricha nova. It has been shown by NMR and x-ray crystallographic analysis that it is capable to form a dimeric quadruplex structure and that a variety of cations, namely K+, Na+, and NH4+, are able to interact with this complex with different affinity, leading to complexes characterized by different local conformations. Thus, in order to improve the knowledge of this kind of molecule, and in particular to provide further insight into the role of monovalent cations in the G-quadruplex folding and conformation, we have investigated by 1H-NMR the effect of the addition of Rb+ and Cs+ to the quadruplex formed by the oligonucleotide d(GGGGTTTTGGGG). [ABSTRACT FROM AUTHOR]

The interest in DNA quadruplex structures has been fueled by the recognition that telomeres, the 3' single stranded guanine-rich overhangs found at the termini of chromosomes, are likely to form G-tetrads type structures important in cell senescence and cancer. In addition to their presence in telomeres, where they may play a role in maintaining the stability and integrity of chromosomes, guanine-rich regions are found in other region of the genome, amongst these is intron 6 of hTERT a gene codifying for the enzyme telomerase. Interestingly, the formation of G-quadruplexes in this region is involved in the down-regulation of telomerase activity caused by an alteration of the hTERT splicing pattern. Therefore, we have analyzed several sequences of that intron by 1H-NMR and CD spectroscopy, and we have found that the sequence d(GGGGTGAAAGGGG) is able to fold in a single well-defined antiparallel quadruplex structure consisting of four G-tetrads, possessing a twofold symmetry, and containing four Gs in a syn glycosidic conformation. [ABSTRACT FROM AUTHOR]

Recently we reported a preliminarry study on the structure of two novel quadruplex structures, Q33 and Q55, formed by the oligodeoxynucleotides 5'TGG3'-3'GGT5' and 3'TGG5'-5'GGT3', respectively. Here we report their solution structures at the atomic level. The obtained structures reveal that Q55 and Q33 possess a different stacking among G-quartets and different twist angle (and therefore different helical winding) at the inversion of polarity level. [ABSTRACT FROM AUTHOR]

Differential scanning calorimetry (DSC) and circular dichroism (CD) techniques were used to investigate the physico-chemical properties of the quadruplexes formed by the two different truncations of human telomeric sequence d(TAGGGT) and d(AGGGT), where the adenines were substituted by 2'-deoxy-8-(hydroxyl)adenosine (A → AOH). CD spectra show that the modified sequences are able to form parallel-stranded quadruplex structure. Analysis of the thermodynamic parameters reveals that the introduction of the modified adenine affects in different way the thermal stability of the [d(TAGGGT)]4 and [d(AGGGT)]4 quadruplexes. [ABSTRACT FROM AUTHOR]

A physico-chemical characterization, based on NMR and CD spectroscopy, of quadruplexes formed by the oligonucleotide d(TGGGT), where two or three Gs are substituted by 8-bromo-2'-deoxyguanosine residues (dGBr), is reported. The oligonucleotidic sequences d(TGBrGBrGT), d(TGBrGGBrT), d(TGGBrGBrT), and d(TGBrGBrGBrT) have been synthesized. Only sequences d(TGBrGGBrT) and d(TGBrGBrGT) were able to fold into a well defined quadruplex structure, and their CD profiles and thermal stabilities turned out to be very different from those observed for the natural counterpart, indicating that the 8-Br-dG residues dramatically affect the structure of the quadruplex. [ABSTRACT FROM AUTHOR]

This work studies the binding properties of distamycin and its carbamoyl analog, containing four pyrrole units, with the [d(TGGGGT)]4 quadruplex by means of isothermal titration calorimetry (ITC). Analysis of the ITC data reveals that drug/quadruplex binding stoichiometry is 1:1 for both interactions and that distamycin analog gives approximately a 10-fold increase in the quadruplex affinity. [ABSTRACT FROM AUTHOR]

The synthesis and preliminary structural studies of ODNs A 8Me GGGT and TA 8Me GGGT, where A 8Me represents 2′-deoxy-8-methyladenosine, are reported. [ABSTRACT FROM AUTHOR]

The effects of incorporation of 8-oxoadenosine in two different truncations of human telomeric sequence forming quadruplex structures are reported. In order to characterise their structures, a combination of NMR and UV spectroscopy and computational techniques were used. Both oligonucleotides have been found to form fourfold symmetric quadruplex structures. As a tautomeric equilibrium between keto and enol forms of 8-oxoadenosine may establish in solution and intrinsic stabilities effects, such as internal H-bonds, for example, may determine the predominance of some particular tautomer, molecular modelling studies were performed on quadruplex structures containing both the tautomeric forms. Both molecules resulted to be thermally less stable than the natural. [ABSTRACT FROM AUTHOR]

Truncated sequences of human telomeric DNA can readily assemble to form parallel stranded quadruplexes containing A- and G-tetrads. The formation of an A-tetrad is highly context-dependent and the relationship between the formation of an A-tetrad and the glycosidic torsion angle of the adenosine residues implicated has not been completely clarified so far. In order to give a further insight in this issue we synthesized the modified oligomers d(ABrGGGT) and d(TABrGGGT), two different truncations of the human telomeric sequence containing a 8-bromoadenosine residue, named ABr. NMR data show that both the modified oligomers are able to perfectly fold into highly symmetric quadruplexes with all strands parallel to each other. Molecular modeling studies were performed on both [d(ABrGGGT)] 4 and [d(TABrGGGT)] 4 , indicating that a bulky substituent, such as a bromine atom at the C8 position of adenines, can force the glycosidic bond to adopt a syn conformation, stabilizing the resulting quadruplexes. [ABSTRACT FROM AUTHOR]

LNAs (locked nucleic acids) are new DNA analogues with higher binding affinities toward nucleic acids than the canonical counterparts mainly due to the characteristic conformational restriction arising from the 2′-O, 4′-C methylene bridge. In light of the promising therapeutic applications and considering the advantageous characteristics of LNAs, such as their high water solubility, easy handling, and synthetic accessibility through the conventional phosphoramidite chemistry, we undertook a study concerning the capability of these nucleic acid analogues to form quadruplex structures. Particularly, we have been investigating the LNA/DNA chimeras corresponding to the well-known DNA sequences 5′-GGTTGGTGTGGTTGG-3′, capable of forming an unimolecular quadruplex. This article deals with the study of the sequence 5′-ggTTggTGTggTTgg-3′ (upper and lower case letters represent DNA and LNA residues, respectively), which, according to CD spectroscopy, is able to fold into a quadruplex structure. [ABSTRACT FROM AUTHOR]

Isothermal titration calorimetry (ITC) is a sensitive technique for probing bimolecular processes and can provide direct information about the binding affinity and stoichiometry and the key thermodynamic parameters involved. ITC has been used to investigate the interaction of the ligand H 2 TMPyP to the two DNA quadruplexes, [d(AGGGT)] 4 and [d(TGGGGT)] 4 . Analysis of the ITC data reveals that porphyrin/quadruplex binding stoichiometry under saturating conditions is 1:2 for [d(AGGGT)] 4 and 2:1 for [d(TGGGGT)] 4 , respectively. [ABSTRACT FROM AUTHOR]

The aim of this work is to compare the physicochemical properties of three oligonucleotidic sequences, d(TGGGT), d(TGGGGT) and d(TGGGGGT), which assemble to form quadruplex structures with the same molecularity, but containing three, four, and five G-quartets, respectively. The addition of one or two G-tetrads greatly increases both the enthalpy and T m values of the quadruplex dissociation. [ABSTRACT FROM AUTHOR]

A NMR structural study of quadruplex [d(TGGGT)]4 containing a modified thymine is reported. The three dimensional structure of the complex is very similar to those of other parallel stranded quadruplexes. The modified thymines (T*) are able, at least in the minimised structures, to form a tetrad containing extra H-bonds through the hydroxyl groups. Nevertheless, in this new tetrad the modified thymines are slightly open towards the solvent respect to the unmodified T-tetrad. [ABSTRACT FROM AUTHOR]

1H-NMR, CD, and UV spectroscopy have been used to investigate the structure of PNA/DNA chimeras forming quadruplex structures. In particular, we synthesized 5′TGGG3′-t (1) and 5′TGG3′-gt (2), where lower and upper case letters indicate PNA and DNA residues, respectively. CD spectrum and all NMR data of (1) are typical of quadruplexes involving four parallel strands. UV melting profile of (1) indicates that its thermal stability is quite similar to that observed for the reference structure [d(TGGGT)]4. 1H-NMR spectrum for 5′TGG3′-gt (2) shows that this oligonucleotide is not able to fold into a single, well-defined species. [ABSTRACT FROM AUTHOR]

Homonuclear NMR techniques have been used to investigate the interactions of the minor groove binding agents distamycin A (Dist-A) and the related drug netropsin (Net) with three quadruplexes characterized by different groove widths: [d(TGGGGT)]4 (Q1), [d(GGGGTTTTGGGG)]2 (Q2), and d(GGGGTTGGGGTGTGGGGTTGGGG) (Q3). Netropsin has been found to be in a fast chemical exchange with all three kinds of quadruplexes, whereas Dist-A interacts tightly with Q1 and, at a less extent, with Q2. In order to determine the degree of selectivity of Dist-A for two- rather than four-stranded DNA, we titrated with Dist-A an equimolar solution of Q1 and the duplex d(CGCAAATTTGCG)2 (D). This comparative 1H-NMR study allowed us to conclude that Dist-A and, consequently, Net possess higher affinity for duplex DNA. [ABSTRACT FROM AUTHOR]

Herein we report the quantification of purine lesions arising from gamma-radiation sourced hydroxyl radicals (HO•) on tertiary dsDNA helical forms of supercoiled (SC), open circular (OC), and linear (L) conformation, along with single-stranded folded and non-folded sequences of guanine-rich DNA in selected G-quadruplex structures. We identify that DNA helical topology and folding plays major, and unexpected, roles in the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) and 8-oxo-7,8-dihydro-2'-deoxyadenosine (8-oxo-dA), along with tandem-type purine lesions 5′,8-cyclo-2′-deoxyguanosine (5′,8-cdG) and 5′,8-cyclo-2′-deoxyadenosine (5′,8-cdA). SC, OC, and L dsDNA conformers together with folded and non-folded G-quadruplexes d[TGGGGT]4(TG4T), d[AGGG(TTAGGG)3] (Tel22), and the mutated tel24 d[TTGGG(TTAGGG)3A] (mutTel24) were exposed to HO•radicals and purine lesions were then quantified via stable isotope dilution LC-MS/MS analysis. Purine oxidation in dsDNA follows L > OC ≫ SC indicating greater damage towards the extended B-DNA topology. Conversely, G-quadruplex sequences were significantly more resistant toward purine oxidation in their unfolded states as compared with G-tetrad folded topologies; this effect is confirmed upon comparative analysis of Tel22 (∼50% solution folded) and mutTel24 (∼90% solution folded). In an effort to identify the accessibly of hydroxyl radicals to quadruplex purine nucleobases, G-quadruplex solvent cavities were then modeled at 1.33 Å with evidence suggesting that folded G-tetrads may act as potential oxidant traps to protect against chromosomal DNA damage. [ABSTRACT FROM AUTHOR]