Quadruplex structure of d(G3T4G3) stabilized by K+ or Na+ is an asymmetric hairpin dimer.

Abstract

The ends of chromosomes contain repeats of guanine-rich sequences that can assume highly compact conformations and are presumed necessary for their biological role in chromosomal stabilization and association. We have investigated the conformational behavior of d(G3T4G3) as a function of the addition of either KCI or NaCl, in the concentration range of 50-200 mM, by using a spectrum of physical techniques and conclude that these salts induce a quadruplex species composed of two strands, each in a hairpin conformation. When salt is added, a large positive signal appears near 290 nm in the CD spectra. UV thermal denaturation curves show a single concentration-dependent transition and provide data for quantitating the thermodynamics of quadruplex formation. In electrophoresis experiments, the quadruplex structure migrates as a single species and more rapidly than the unstructured single strand. NMR spectra in the presence of KCI or NaCl indicate that the structure formed is asymmetric. Equilibrium ultracentrifugation studies confirm that these quadruplexes are composed of two strands of d(G3T4G3). Possible models for this structure are discussed.The ends of chromosomes, termed telomeres, contain a single-stranded 3' overhang composed of repeats of guaninerich sequences, such as d(G4T4) and d(G4T2). Telomeric DNA sequences can take on highly compact conformations that are presumed necessary for their biological role in chromosomal stabilization and association (1-3). Such structures often entail hairpin or fold-back forms involving one or two strands of G-rich DNA (4). Hastie and Allshire (5) have discussed the possible role of quadruplex structures in recombination, and Sen and Gilbert (6) have described four-stranded guaninerich structures (G4 DNA) formed by several telomeric sequences. Furthermore, there is evidence that G-rich runs in immunoglobulin switch regions may form four-stranded quadruplex structures (7). Guanine-rich sequences have also been found in gene promoter regions (8,9).In all of these unusual forms, the fundamental structural unit-the guanine quartet-is composed of four guanines arrayed in a square planar configuration. Each guanine interacts with each of the two adjacent guanines through two Hoogsteen-like hydrogen bonds. The simplest system potentially capable of guanine quartet formation consists of guanine nucleosides or nucleotides, solutions of which have long been known to form gels (10). Recently, Lipanov et al. (11) have obtained crystals of Na25'-GMP. Preliminary x-ray analysis has revealed a helical pattern that can be interpreted in terms of a quadruple helix made up of three guanine quartets. Earlier reports have also appeared describing parallel four-stranded structures for poly(G) and poly(I) (12). In the last 5 years, several telomeric sequences have been studied using physical and structural methods (13-18). Henderson et al. (15) have examined a variety of sequences with enhanced electrophoretic mobility that exhibited low-field exchangeable protons in the NMR spectra,...

G-Quadruplex Stabilizer 3,6-Bis(1-Methyl-4-Vinylpyridinium)Carbazole Diiodide Induces Accelerated Senescence and Inhibits Tumorigenic Properties in Cancer Cells

“…To evaluate the effect of BMVC to the G-quadruplex DNA of human telomeric sequences, d(T 2 AG 3 ) 4 (Hum), we applied circular dichroism to monitor the melting temperature (Tm) of Hum quadruplexes. It has been documented that parallel fourstranded quadruplexes give a positive band f260 nm and a negative band f240 nm (27), whereas antiparallel folded quadruplexes have two positive bands around 245 and 290 nm and a negative band f265 nm (28). The 295-nm circular dichroism band was measured as a function of temperature to determine the Tm of Hum24 quadruplexes in the presence of carbazole and BMVC (29).…”

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

“…NMR spectroscopy showed that each of the DNA strands self-assembled into individual bimolecular quadruplexes, with distinct topologies, in the presence of monovalent cations [21][22][23][24][25][26][27][28]. The monovalent cations are necessary to coordinate in the core of the quaduplexes and so to stabilize the structures.…”

“…The monovalent cations are necessary to coordinate in the core of the quaduplexes and so to stabilize the structures. [Note: Subsequently in this paper, the sequences will be described as d( 2 by the absence of a middle G-quartet (Figure 1d), so that the guanine bases are arranged 5=-syn-synanti-loop-syn-anti-anti-3= in one strand and 5=-syn-antianti-loop-syn-syn-anti-3= in the other [23][24][25][26]. In contrast, the structure of the bimolecular [d(G 3 G 4 )] 2 is special (Figure 1c), in that one guanine residue leaps over the middle G-quartet, turning an antiparallel strand parallel to its own half, and so that two dangling guanines reside at opposite ends of the quadruplex [27,28].…”

“…The guanine bases in each strand of this alternate syn-anti glycosidic torsion angles along the stacking quartets [29]. NMR studies show that these bimolecular quadruplexes retain the same general folds in Na ϩ , K ϩ , or NH 4 ϩ solution ( Figure 1b-d) [22,24,25,28], except for the strand d(G 4 G 3 ), which folds into the bimolecular structure (Figure 1e) only in Na ϩ solution, but forms multiple structures in the presence of K ϩ and NH 4 ϩ [29]. Using ESI-MS, we expect to detect specific interactions between strand and strand, as well as between quadruplex and monovalent ammonium ion.…”

Electrostatics dominate quadruplex stability

“…For example, the sequence d(G 3 T 4 G 3 ) forms a two-stranded quadruplex in the presence of excess amounts of either Na þ or K þ . [33][34][35] In contrast to the unimolecular quadruplexes, addition of near stoichiometric amounts of Ba 2þ , for example, results in virtually no quadruplex formation, as illustrated in Figure 2. However, if methanol is present, then a peak near 300 nm, indicative of antiparallel quadruplex formation, appears.…”