Multiplex formation of repetitive sequences d([G]nA)m through non-WC pairings: crystal structure of d(gcGAGGGAgc).

The crystal structure of a DNA fragment with the sequence d(gcGA[G](3)Agc), a shortened and mutated derivative of the VNTR sequence d(ccGA[G](4)Agg), has been determined by X-ray analyses at 1.8 A resolution to investigate structures and mechanisms of a non-coding DNA. This sequence forms an antiparallel duplex with two WC pairings at the both ends and six consecutive non-WC pairings in the major part.

Octaplex formation of d(GCGAAAGC) with the double A-quartet.

We reported that the sequence gcGA[X]1Agc adopts a specific structure to form several multiplexes. In the case of X=G, an octaplex formation occurs, in which the stacked double G-quartet stabilizes the architecture through potassium cation mediations. In the case of a mutant X=A, too, it has been found that the oligomers form an octaplex.

Structural stability and length of I-motif of G-quartet in DNA octaplex.

To confirm that longer DNA strands with sequences gcGA[G](n)Agc can form an octaplex, X-ray analyses at n>1 have been performed. Several crystals of d(gcGA[G]2Agc) are obtained under different conditions. It has been found that the crystal obtained with cobalt hexamine suggests a hexaplex formation, while the crystal obtained with Mg2+ and Na+ ions suggests an octaplex formation.

DNA octaplex formation with an I-motif of water-mediated A-quartets: reinterpretation of the crystal structure of d(GCGAAAGC).

The crystal structure of the tetragonal form of d(gcGAAAgc) has been revised and reasonably refined including the disordered residues. The two DNA strands form a base-intercalated duplex, and the four duplexes are assembled according to the crystallographic 222 symmetry to form an octaplex. In the central region, the eight strands are associated by I-motif of double A-quartets.

Crystal structure of d(gcGXGAgc) with X=G: a mutation at X is possible to occur in a base-intercalated duplex for multiplex formation.

DNA fragments with the sequences d(gcGX[Y]n Agc) (n=1, X=A, and Y=A, T, or G)form base-intercalated duplexes, which is a basic unit for formation of multiplexes such as octaplex and hexaplex. To examine the stability of multiplexes, a DNA with X=Y=G and n=1 was crystallized under conditions different from those of the previously determined sequences, and its crystal structure has been determined. The two strands are coupled in an anti-parallel fashion to form a base-intercalated duplex, in which the first and second residues form Watson-Crick type G:C pairs and the third and sixth residues form a sheared G:A pairs at both ends of the duplex.