DNA nanotechnology excels at rationally designing bottom-up structures that can functionally replicate naturally occurring proteins. Here we describe the design and generation of a stable DNA-based nanopore that structurally mimics the amphiphilic nature of protein pores and inserts into bilayers to support a steady transmembrane flow of ions. The pore carries an outer hydrophobic belt comprised of small chemical alkyl groups which mask the negatively charged oligonucleotide backbone. This modification overcomes the otherwise inherent energetic mismatch to the hydrophobic environment of the membrane. By merging the fields of nanopores and DNA nanotechnology, we expect that the small membrane-spanning DNA pore will help open up the design of entirely new molecular devices for a broad range of applications including sensing, electric circuits, catalysis, and research into nanofluidics and controlled transmembrane transport.

Holding tight: An artificial membrane nanopore assembled from DNA oligonucleotides carries porphyrin tags (red), which anchor the nanostructure into the lipid bilayer. The porphyrin moieties also act as fluorescent dyes to aid the microscopic visualization of the DNA nanopore.

Tetraphenyl porphyrin substituted deoxyuridine was used as a building block to create discrete multiporphyrin arrays via site specific incorporation into DNA. The successful covalent attachment of up to 11 tetraphenyl porphyrins in a row onto DNA shows that there is virtually no limitation in the amount of substituents, and the porphyrin arrays thus obtained reach the nanometer scale (approximately 10 nm). The porphyrin substituents are located in the major groove of the dsDNA and destabilize the duplex by deltaT(m) 5-7 degrees C per porphyrin modification. Force-field structure minimization shows that the porphyrins are either in-line with the groove in isolated modifications or aligned parallel to the nucleobases in adjacent modifications. The CD signals of the porphyrins are dominated by a negative peak arising from the intrinsic properties of the building block. In the single strands, the porphyrins induce stabilization of a secondary helical structure which is confined to the porphyrin modified part. This arrangement can be reproduced by force-field minimization and reveals an elongated helical arrangement compared to the double helix of the porphyrin-DNA. This secondary structure is disrupted above approximately 55 degrees C (T(p)) which is shown by various melting experiments. Both absorption and emission spectroscopy disclose electronic interactions between the porphyrin units upon stacking along the outer rim of the DNA leading to a broadening of the absorbance and a quenching of the emission. The single-stranded and double-stranded form show different spectroscopic properties due to the different arrangement of the porphyrins. Above T(p) the electronic properties (absorption and emission) of the porphyrins change compared to room temperature measurements due to the disruption of the porphyrin stacking at high temperature. The covalent attachment of porphyrins to DNA is therefore a suitable way of creating helical stacks of porphyrins on the nanometer scale.

Oligonucleotides have recently gained increased attraction as a supramolecular scaffold for the design and synthesis of functional molecules on the nanometre scale. This tutorial review focuses on the recent progress in this highly active field of research with an emphasis on covalent modifications of DNA; non-covalent interactions of DNA with molecules such as groove binders or intercalators are not part of this review. Both terminal and internal modifications are covered, and the various points of attachment (nucleobase, sugar moiety or phosphodiester backbone) are compared. Using selected examples of the recent literature, the diversity of the functionalities that have been incorporated into DNA strands is discussed.