A DNA pentaplex incorporating nucleobase quintets

Abstract

Supramolecular self-assembly is an integral step in the formation of many biological structures. Here we report a DNA pentaplex that derives from a metal-assisted, hydrogen bond-mediated self-assembly process. In particular, cesium ions are found to induce pentameric assembly of DNA bearing the nonstandard nucleobase iso-guanine. The pentaplex was designed by using a simple algorithm to predict nucleobase structural requirements within a quintet motif. The design principles are general and should extend to complexes beyond pentaplex. Structures exhibiting molecularities of five or more were previously accessible to peptides, but not nucleic acids.

Figure 1

Working design for nucleobase motifs leading to expanded DNA molecularities. (a, from left to right) Schematic representation of a circularly modular quartet motif with ideal sector angle of 90°, ab initio optimized geometry of a G-quartet motif, and actual 90° sector angle for G defined by the intersection of vectors along the van der Waals donor-donor and acceptor-acceptor atom surfaces using the geometry from its optimized quartet. (b, from left to right) Schematic representation of a circularly modular quintet motif with ideal sector angle of 72°, ab initio optimized geometry of an iG-quintet motif, and actual 67° sector angle for iG defined by the intersection of vectors along the van der Waals donor-donor and acceptor-acceptor atom surfaces using the geometry from its optimized quintet. (c) Schematic representation of a circularly modular n-mer motif with ideal sector angle of 360°/n.

Figure 2

Autoradiograms of native 20% polyacrylamide gels demonstrating the number of electrophoretic bands that emanate from complexes formed by two different lengths of iG₄-bearing DNA strands. Complexes were formed by incubating DNA strands separately or together in the presence of K⁺ and/or Cs⁺, as indicated at the top of each lane.

Figure 3

Schematic representation of electrophoretically distinguishable tetraplexes and pentaplexes that may result when two different lengths of DNA strands are combined. (a) Five parallel-stranded tetraplexes that may be distinguished; (Lower) strand ratios that were experimentally determined. (b) Six parallel-stranded pentaplexes that may be distinguished; (Lower) experimentally determined strand ratios.

Figure 4

Autoradiograms of denaturing polyacrylamide gels showing the result of UV photocrosslinking experiments with different lengths of iG₄-bearing oligonucleotides in K⁺ or Cs⁺. (a) ³²P-end-labeled oligonucleotides were incubated either alone, or combined together, in 0.5 M of KCl at 4°C for 2 days as indicated at the top of each lane. In “crossover” control lanes 4 and 5, a ³²P-end-labeled single-strand “challenge oligomer” was added after the 2 days of incubation, just before irradiation, as indicated at the top of these lanes. Samples then were irradiated with UV light for 15 min at 4°C. Products of UV irradiation were electrophoresed on a 20% denaturing (7 M urea) polyacrylamide gel at 45°C with 1× TBE (Tris/boric acid/EDTA, pH 8.6) as running buffer. (b) The same procedure was followed as detailed for a except CsCl was used in place of KCl. Labels located in between the panels in all capital letters describe products of T₈iG₄T, labels in all lowercase letters refer to those of T₄iG₄T, and “mixed” refers to irradiation products having elements derived from both of these oligomers.

Figure 5

Pentaplex of iG₄ DNA strands incorporating two cesium ions. The structure shown results from unrestrained minimization of the complex by using the amber* forcefield implemented in Macromodel. Two cesium ions are incorporated into the model because a three-ion model with an ion between each layer proved unstable. The average intrastrand phosphate-phosphate distance is observed to be 6.8 Å.

Figure 6

Hartree-Fock optimized nucleobase quintet-metal cation geometries. (a) iG-quintet/Cs⁺/iG-quintet 3–21 optimized geometry; the Cs⁺ is positioned between quintet layers. (b) iG-quintet/K⁺ 3–21G* optimized geometry; the K⁺ lies in the quintet plane.