G-quadruplexes in telomeric repeats are conserved in a solvent-free environment

Four individual quadruplexes, which are self-assembled in ammonium acetate solution from telomeric sequences of closely related DNA strands-d(G 4 T 4 G 4 ), d(G 3 T 4 G 4 ), d(G 3 T 4 G 3 ), and d(G 4 T 4 G 3 )-have been detected in the gas phase using electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR-MS). The bimolecular quadruplexes associate with the same number of NH 4 ϩ in the gas phase as NMR shows that they do in solution. The quadruplex structures formed in solution are maintained in the gas phase. Furthermore, the mass spectra show that the bimolecular quadruplexes generated by the strands d( Of particular interest is a telomeric DNA that consists of repeating sequences, such as TTAGGG in vertebrates and TTTTGGGG in Oxytricha. These sequences have been proven to fold into quadruplexes in vitro and potentially have the function of inhibiting telomerase activities. The telomerase, a reverse enzyme expressed in more than 80% of tumor cells, responds for the replication of the telomeric DNA and has been considered as a target for anticancer drug design [4,5]. Guanine-rich oligonucleotides can also self-associate into higher-order structures, such as supermolecular G-wires [6], which serve as artificial ion channels and ion receptors and which have a wide range of applications in medicine, materials science, and nanotechnology [2,7,8].DNA quadruplex structures can be formed from one, two, or four oligonucleotide strands and are stabilized by coordinated monovalent cations acting between adjacent G-quartets [1, 9 -11]. The strands with single repeating guanine sequences usually arrange themselves from the 5= to the 3= end in parallel four-stranded quadruplexes, whereas the strands with more than two tracks of repeating guanine units can form a number of quadruplexes that are distinguished by their strand number, strand orientation, and loop types. The structural topologies of DNA quadruplexes also drastically change with slight changes of the DNA sequences, with the nature of the cationic species and with their concentrations in solution. As a result, the structures of DNA quadruplexes are far from being fully recognized and have been of concern to researchers because of their potential implications in biological and structural applications.Development in electrospray ionization mass spectrometry (ESI-MS) has been shown to be superior in the analysis of DNA noncovalent complexes in terms of speed, sensitivity, stoichiometry, and specificity compared to other analytical techniques [12]. Previous work by ourselves and others have shown that some specific DNA conformations, such as hairpins, triplexes, and quadruplexes formed in solution, can survive in the gas phase [13][14][15][16][17][18][19][20]. This prompted us to explore the structural properties of DNA quadruplexes with ESI-MS so as to add new information concerning them.

Section: Detection Of the Bimolecular Dna Quadruplexesmentioning

confidence: 99%

“…Previous work by ourselves and others have shown that some specific DNA conformations, such as hairpins, triplexes, and quadruplexes formed in solution, can survive in the gas phase [13][14][15][16][17][18][19][20]. This prompted us to explore the structural properties of DNA quadruplexes with ESI-MS so as to add new information concerning them.…”

mentioning

confidence: 99%

Bimolecular quadruplexes and their transitions to higher-order molecular structures detected by ESI-FTICR-MS

Guo

Liu

2007

“…Measuring the drift time directly linked to the mobility of an ion yields information about its structure because compact ions, which have necessarily relatively small collision cross sections, move faster than extended ones, which have large collision cross sections. The gas-phase conformations of many oligonucleotides have been investigated by ion mobility: deprotonated mononucleotides [19,20], dinucleotides [21], trinucleotides [22], deprotonated oligothymidines [23], DNA complexes [24], G-quadruplex [25,26], etc.…”

mentioning

confidence: 99%

Detection of oligonucleotide gas-phase conformers: H/D exchange and ion mobility as complementary techniques

Balbeur

Widart

Leyh

et al. 2008

Self Cite

Gas-phase hydrogen/deuterium exchange of small oligonucleotides (dTG, dC 6 and C 6 ) with CD 3 0D was performed in the second hexapole of a Fourier transform ion-cyclotron resonance (FTICR) mass spectrometer. Ion activation experiments were conducted by accelerating the ions at the entrance of the H/D exchange cell under conditions promoting exclusively collisional isomerization. These experiments allowed us to assess the presence of several conformers, and to probe the height of the isomerization barrier separating these conformers. Hydrogen/deuterium (H/D) exchange combined with mass spectrometry has become an efficient tool for obtaining conformational information about isolated biomolecules in the gas phase [1-18J. The rate of gasphase H/D exchange has been shown to be a function of the reagent used for exchange (nature and concentration), the charge state of the biomolecule, the gasphase basicity/acidity of exchangeable sites, and the internal structure of the biomolecular ions [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18]. Probing the number of exchanged hydrogens and the H/D exchange kinetics makes it possible to distinguish ion populations and to elucidate their structural and thermochemical features. Gas-phase H/D exchange has been applied extensively for structural studies of proteins and peptides [1-6J and is now also most commonly used for structural characterizations of oligonucleotides [7][8][9][10][11][12][13][14][15][16][17][18]. Mononucleotides studies have shown Address reprint requests to Miss Dorothee Balbeur, Laboratory of Mass Spectrometry, University of Liege, 3 Allee de la Chimie Bat. B6c, B-4000 Liege, Belgium. E-mail: dbalbeur@ulg.ac.be that the extent and rate of exchange depend on the nucleobase (nature, space orientation, and gas-phase acidity of the exchangeable hydrogen), and on the position (3' or 5') and flexibility of the terminal phosphate group [7-11J. Moreover, these studies have been in favor of the relay mechanism [7][8][9][10][11]. The latter was shown not to be relevant for dinucleotides [14J. Furthermore, gas-phase H/D exchange of dinucleotides has been shown to be controlled by hydrogen accessibility rather than by the chemical nature of the heteroatom bearing the exchangeable hydrogen. Studies of larger oligonucleotides have also been published. Gas-phase H/D exchange reactions were performed on DNA duplexes and on the corresponding single strands. They showed that H/D exchange was slower for the duplex than for the single strands [15, 16J. In a previous study, a quadruplex was shown to undergo very fast H/D exchange compared with its constituent monomer, an unexpected situation for such a compact and rigid structure [17J. Gas-phase H/D exchange of 5-mer phosphorothioate oligonucleotides was also examined; bimodal exchange profiles were observed, suggesting that each ion could adopt more than one gas-phase conformation [18J.The present work focuses on the bimodal H/D exchange of oligonucleotides. A simple interpretation of such a behavior is that one ...

“…For example, the dissociation profiles of a series of 6-, 8-, and 12-base pair selfcomplementary duplexes have been shown to correspond to the known solution melting behavior of DNA, demonstrating the utility of CAD to provide relative comparisons of the gas-phase stabilities of DNA duplexes [30]. Fewer studies have been undertaken to elucidate the gas-phase structures or stabilities of quadruplex DNA [32][33][34][35][36][37]. One recent study focused on using ion mobility mass spectrometry and molecular dynamics simulations to examine gas-phase conformations of quadruplexes with different lengths of the telomeric repeat d(TTAGGG) n where n ϭ 1, 2, 4, and 6 [33].…”

mentioning

confidence: 99%

Gas-phase stability of G-quadruplex DNA determined by electrospray ionization tandem mass spectrometry and molecular dynamics simulations

Mazzitelli

Wang²,

Smith

et al. 2007

) 4 , were investigated using molecular dynamics simulations and electrospray ionization mass spectrometry (ESI-MS). MD simulations revealed that the G-quadruplexes maintained their structures in the gas phase although the G-quartets were distorted to some degree and ammonium ions, retained by [d(TG 4 T)] 4 and [d(T 2 G 3 T)] 4 , played a key role in stabilizing the tetrad structure. Energyvariable collisional activated dissociation was used to assess the relative stabilities of each quadruplex based on E 1/2 values, and the resulting order of relative stabilities was found to beThe stabilities from the E 1/2 values generally paralleled the RMSD and relative free energies of the quadruplexes based on the MD energy analysis. One exception to the general agreement is [d(G 4 T 4 G 4 )] 2 , which had the lowest E 1/2 value, but was determined to be the most stable quadruplex according to the free-energy analysis and ranked fourth based on the RMSD comparison. This discrepancy is attributed to differences in the fragmentation pathway of the quadruplex. T he basis of many anticancer and antitumor therapies is the interaction between small molecule drugs and nucleic acid structures. While most current DNA-interactive therapies target duplex DNA, G-quadruplex DNA has attracted recent interest as a potential anti-cancer drug target because of its role in telomere maintenance [1][2][3]. Telomeres are the regions of non-coding DNA found on the extremities of chromosomes [4]. In addition to protecting the chromosomes from fusion and degradation, telomeres allow for the complete replication of the chromosomal DNA. With each subsequent cell division process, the length of the telomeres is shortened until a critical length is reached, leading to cell senescence and death [5]. Telomerase is a reverse transcriptase enzyme that is responsible for maintaining the length of telomeric DNA. While this enzyme is inactive in most human somatic cells, high levels of telomerase activity are found in 80% to 90% of human cancer cells [6,7].Telomeric DNA is composed of tandem repeats of G-rich sequences, such as the d(T 2 AG 3 ) n sequence in mammals [8], d(T 2 G 4 ) n in Tetrahymena [9], and d(T 4 G 4 ) n in Oxytricha [10]. These and other G-rich sequences have been shown to form G-quadruplex structures in vitro. G-quadruplex DNA forms via Hoogsteen hydrogen bonding between a planar arrangement of four guanine nucleobases, termed a G-quartet. In the quadruplex structure, the G-quartets stack on top of one another and the overall structure is stabilized by monovalent cations such as Na ϩ , K ϩ , or NH 4 ϩ coordinated in the central cavity of the tetrad [11]. Structural polymorphism resulting from different quadruplex sequences and strand orientations has been demonstrated in vitro [12]. A DNA sequence containing a single G-rich repeat can form a four-stranded parallel G-quadruplex. Strands containing two or more G-rich regions can form G-G hairpins and dimerize in different orientations to form a two-stranded quadruplex, while a sequence w...