Caffeine-induced release of small molecules from DNA nanostructures

Abstract

Several planar aromatic molecules are known to intercalate between base pairs of double-stranded DNA. This mode of interaction has been used to stain DNA as well as to load drug molecules onto DNA-based nanostructures. Some small molecules are also known to induce deintercalation in double-stranded DNA, one such molecule being caffeine. Here, we compared the ability of caffeine to cause deintercalation of ethidium bromide, a representative DNA intercalator, from duplex DNA and three DNA motifs of increasing structural complexity (four-way junction, double crossover motif, and DNA tensegrity triangle). We found that caffeine impedes the binding of ethidium bromide in all these structures to the same extent, with some differences in deintercalation profiles. Our results can be useful in the design of DNA nanocarriers for intercalating drugs, where drug release can be chemically stimulated by other small molecules.

Keywords: Chemistry; Molecular self-assembly; Supramolecular chemistry.

Graphical abstract

Figure 1

Design and overview (A) Schematic showing the intercalation of ethidium bromide (EBr) to DNA and deintercalation triggered by caffeine. (B) Model DNA nanostructures used in this study: a duplex, a four-way junction (4WJ), a double crossover (DX) motif, and a 3-helical-turn per edge tensegrity triangle (3TT) motif. (C) Non-denaturing PAGE showing assembly of the structures. See Table S1 for full sequences and Figure S1 for designs of the structures with sequences.

Figure 2

Binding of ethidium bromide to DNA nanostructures Fold change in fluorescence emission at 600 nm of EBr (20–80 μM) upon intercalation into different concentrations of (A) duplex DNA, (B) 4WJ, (C) DX motif, and (D) 3TT. DNA concentrations are indicated as base pairs (BP). Data represent mean and error propagated from standard deviations of triplicate experiments. See Figure S2 for representative fluorescence spectra and Figure S3 for raw fluorescence data.

Figure 3

Caffeine inhibits binding of ethidium bromide to DNA nanostructures Calculated intercalation indices showing reduced binding of EBr (20 μM) to DNA nanostructures (40 μM BP) with increasing concentrations of caffeine. Values are normalized to intensities obtained in the absence of caffeine. Data represent mean and error propagated from standard deviations of triplicate experiments. See Figure S4 for a plot containing data from all four structures overlapped.

Figure 4

Caffeine-induced deintercalation of ethidium bromide from DNA nanostructures (A) Non-denaturing gels for different DNA structures stained with ethidium bromide and soaked either in buffer (top row) or caffeine (bottom row). (B) Percent deintercalation in different DNA nanostructures measured using the decrease in band intensity in buffer and caffeine. Data represent mean and error propagated from standard deviations of triplicate experiments. See Figure S6 for gel images of all time points and Figure S7 for a plot containing data from all four structures overlapped.