Tetraplex structures

Tetraplex structures have also been observed for G-rich repeating sequences associated with the human fragile X syndrome 275/276 This is the most common cause of inherited mental retardation and appears to arise as a result of the presence of an excessive number of repeats of the trinucleotide sequence (CGG)n. For normal persons n = 60 or less for healthy carriers n may be as high as 200 but for sick individuals it may be much higher.275 The structure in solution, as determined by NMR spectroscopy, is shown in Fig. 5-26. Another variant of four-stranded DNA, which arises from cytosine-rich DNA, contains C CH+ pairs such as the following at low pH.277 279... 

It has been recognized that the stabilization by interactions with metal ions is essential in the formation of tetraplex structures. Based on the electrostatic potential analysis of the tetrads, a number of tetrads are suggested as the possible hosts of cations. The interaction between the cation and the tetrads remarkably alter the bonding patterns of the tetrads. 

Wang Y, Patel DJ. 1993. Solution structure of the human telomeric repeat d[AG3(T2AG3)3] G-tetraplex. Structure 1 263-82... 

Y. Wang and D.J. Patel, Solution structure of the Tetrahymena telomeric repeat d(T2G4)4 G-tetraplex, Structure, 1994, 2, 1141-1156. 

B. Pan, Y. Xiong, K. Shi, J. Deng and M. Sundaralingam, Crystal structure of an RNA purine-rich tetraplex containing adenine tetrads Implications for specific binding in RNA tetraplexes, Structure, 2003, 11, 815-823. 

In addition to the discovery of new ligands that interact with tetraplex structures, the DCC concept has also been used by the Balasubramanian, Lehn,... 

Porphyrins are a class of ligands that are more commonly associated with proteins than DNA, but their large stacking surface area has led some groups to explore their ability to bind (intercalatively) to tetraplex structures [327-329]. The binding of the porphyrins to the tetraplexes has been monitored by visible absorption and fluorescence titrations [327-329]. 

Base triads do, of course, occur in nucleic acid triplexes. However, tetraplex structures may also contain triads and in RNA structures triads often play a crucial role. Triplexes are formed by the interaction of a third strand in the major groove of a double helix. The duplex has to be composed of a homopurine-homopyrimidine sequence (piuine - R, pyrimidine - Y). The third strand can bind in a parallel or antiparallel orientation to one of the duplex strands. In parallel orientation, a homopyrimidine third strand binds to the homopurine strand of the duplex (YRY). This leads to the two canonical triads TAT and C+GC. Protonation of C (C+) at N3 is required for the formation of two H-bonds between C and G. Therefore, parallel triplexes are pH dependent. These structures have two canonical base triads TAT and C+GC. For an antiparallel orientation of the third strand relative to the binding duplex strand, a homopruine sequence is required that binds to the homopurine strand of the duplex (RRY). This results in the canonical triads GGC, AAT and TAT, where however the TAT triad is different to the corresponding triad in parallel triplexes. In addition to these standard triads, triplexes can also accommodate non-canonical base triads. Fig. 3 shows the two canonical triads C+GC and TAT in an intra-molecular triplex consisting of a DNA duplex and... 

Much less is known about non-G-tetrads. Even though file telomeric sequences are G-rich, they also include other bases. Therefore, the existence of non-G-tetrads cannot be excluded. Indeed, A-, T- and C-tetrads have recently been foimd in DNA tetraplex structures. Moreover, an interesting U-tetrad linked by C-H...O contacts only occurs in an exceptionally stable RNA tetraplex with G- and U-tetrads. Very recently a high-resolution x-ray structure for the same sequence has become available, that allows for a comparison between the x-ray and NMR results. ... 

In addition to tetrads comprised of just one base type, mixed tetrads have been observed. Approximately planar GCGC tetrads occur in a tetraplex structure with the fragile X syndrome repeat sequence (PDB code la8w) and a tetraplex structure fi om admo-associated viral DNA (PDB code la8n). On the other hand, significantly non-planar GCGC tetrad are also known, see Fig... 

Fig. 7. Non-planar GCGC tetrads in a DNA tetraplex structure of d(GCATGCT) (PDB code 184d). The tetrad adopts the topology IV.1(24).

Fig. 9. Base heptad in a DNA tetraplex structure. The heptad has the toplogy VII.3(332i I3). At the dotted lines the H-bond distances are indicated.

Fig. 10. Base octad in an RNA tetraplex structure (UGGGGU)4 (PDB code IjSg). Two tetraplexes stack on each other. The 5 -end uridines rotate around the backbone and form octads with the neighboring G-tetrad. The octad topology is VIII.3(34l4).

As already noted, metal ions play a vital role in DNA tetraplex structures. The first MD study that did address this point was published in 1994. It was found that the simulation could reproduce a few properties of the structure but could not correctly describe the metal ion nucleic acid interaction. This is not surprising as at that time the particle-mesh Ewald roach was not yet available. Later simulations do not suffer from this deficioicy. 

A nanosecond MD simulation of d(T4G4T4) from 1999 h clearly shown a destabilization of the syston when removing the metal ions fix>m the central ion chatmel. Interestingly, a very stable alternative conformation involving a guanine triad has been found in the simulations. This observation can be related to the fact that experimental DNA tetraplex structures may not only contain tetrads but also triads, pentads, hexads, heptads and octads. The two iimer G-tetrads of the four stacked tetrad planes exhibit contrary to the expoimental structure a bifurcated H-bond pattern. 

As for triplexes the effect of structural modifications has also been studied for DNA-tetraplexes. The simulations on structures containing inosine, 6-thioguanine and 6-thiopurine indicate that a nucleic add stem formed fixnn G-tetrads can easily incorporate inosine without any marked effects on structure and stability. On the other hand, 6-thioguanine and 6-thiopurine have more or less adverse effects. They cause a destabilization of the tetraplex structure... 

In other nanosecond MD simulations on parallel and antiparallel G-tetrads the effect of coordinated cations on structure, flexibility and stability has been studied. A further MD study has started with G-tetraplex structure without any ions in the initial structure. The tetraplex structure is not disrupted but undergoes small structural changes, which allow the Na ions to move into the empty central ion channel. Even though not all potential ion binding sites were occupied during the time of simulation the structure remained stable. 

We would like to thank our co-workers M. Brandi and C. Schneider who were involved in our computational studies on nucleic acids. J. Reichert has helped with the topologies. We are grateful to M. Katahira who provided the coordinates of a hq)tad-containmg DNA tetraplex structure prior to PDB deposition. The Thtiringer Ministerixim fur Wissenschaft, Forschung und Kunst has funded part of the work. 

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