Switchable enzyme/DNAzyme cascades by the reconfiguration of DNA nanostructures.

Mimicking cellular transformations and signal transduction pathways by means of biocatalytic cascades proceeding in organized media is a scientific challenge. We describe two DNA machines that enable the "ON/OFF" switchable activation and deactivation of three-component biocatalytic cascades. One system consists of a reconfigurable DNA tweezers-type structure, whereas in the second system the catalytic cascade proceeds on a switchable DNA clamp scaffold. The three-component catalytic cascades consist of ß-galactosidase (ß-Gal), glucose oxidase (GOx), and the K(+) -ion-stabilized hemin-G-quadruplex horseradish peroxidase (HRP)-mimicking DNAzyme. The hemin-G-quadruplex-bridged closed structure of the tweezers or clamp allows the biocatalytic cascades to operate (switched "ON''), whereas separation of the hemin-G-quadruplex by means of 18-crown-6-ether opens the tweezers/clamp structures, thus blocking the catalytic cascade (switched "OFF"). This study is complemented by two-component, switchable biocatalytic cascades composed of GOx and hemin-G-quadruplex assembled on hairpin-bridged DNA tweezers or clamp nanostructures.

Nucleic acid/quantum dots (QDs) hybrid systems for optical and photoelectrochemical sensing.

Nucleic acid/semiconductor quantum dots (QDs) hybrid systems combine the recognition and catalytic properties of nucleic acids with the unique photophysical features of QDs. These functions of nucleic acid/QDs hybrids are implemented to develop different optical sensing platforms for the detection of DNA, aptamer-substrate complexes, and metal ions. Different photophysical mechanisms including fluorescence, electron transfer quenching, fluorescence resonance energy transfer (FRET), and chemiluminescence resonance energy transfer (CRET) are used to develop the sensor systems. The size-controlled luminescence properties of QDs are further implemented for the multiplexed, parallel analysis of several DNAs, aptamer-substrate complexes, or mixtures of ions. Similarly, methods to amplify the sensing events through the biocatalytic regeneration of the analyte were developed. An additional paradigm in the implementation of nucleic acid/QDs hybrids for sensing applications involves the integration of the systems with electrodes, and the generation of photocurrents as transduction signals for the sensing events. Finally, semiconductor QDs conjugated to functional DNA machines, such as "walker" systems, provide an effective optical label for probing the dynamics and mechanical functions of the molecular devices. The present article addresses the recent advances in the application of nucleic acid/QDs hybrids for sensing applications and DNA nanotechnology, and discusses future perspectives of these hybrid materials.

Optical aptasensors for the analysis of the vascular endothelial growth factor (VEGF).

The vascular endothelial growth factor, VEGF, is an important biomarker for different diseases and clinical disorders. We present a series of optical aptasensor-based sensing platforms for VEGF that include the following: (i) A FRET-based sensor that involves the VEGF-induced separation of aptamer-functionalized quantum dots blocked by a quencher nucleic acid (detection limit 1 nM). (ii) A FRET-based sensor based on the VEGF-induced assembly of the aptamer subunits functionalized with QDs and a dye acceptor (Cy5), respectively (detection limit 12 nM). (iii) A chemiluminescence aptasensor based on VEGF-induced assembly of a hemin/G-quadruplex catalyst (detection limit 18 nM). (iv) A chemiluminescence aptasensor based on the VEGF-stimulated assembly of two aptamer subunits into the hemin/G-quadruplex catalyst (detection limit 2.6 nM). (v) A chemiluminescence resonance energy transfer (CRET) aptasensor based on the VEGF-induced assembly of a semiconductor QDs-hemin/G-quadruplex supramolecular structure (detection limit 875 pM). Furthermore, an amplified optical aptasensor system based on the Exonuclease III (Exo III) recycling of the VEGF analyte was developed. In this system, one aptamer subunit is modified at its 5' and 3' ends with QDs and a black hole quencher, respectively. The VEGF-induced self-assembly of the aptamer subunits result in the digestion of the quencher units and the autonomous recycling of the analyte, while triggering-on the luminescence of the QDs (detection limit 5 pM). The system was implemented to analyze VEGF in human sera samples.

Following glucose oxidase activity by chemiluminescence and chemiluminescence resonance energy transfer (CRET) processes involving enzyme-DNAzyme conjugates.

A hybrid consisting of glucose oxidase-functionalized with hemin/G-quadruplex units is used for the chemiluminescence detection of glucose. The glucose oxidase-mediated oxidation of glucose yields gluconic acid and H(2)O(2). The latter in the presence of luminol acts as substrate for the hemin/G-quadruplex-catalyzed generation of chemiluminescence. The glucose oxidase/hemin G-quadruplex hybrid was immobilized on CdSe/ZnS quantum dots (QDs). The light generated by the hybrid, in the presence of glucose, activated a chemiluminescence resonance energy transfer process to the QDs, resulting in the luminescence of the QDs. The intensities of the luminescence of the QDs at different concentrations of glucose provided an optical means to detect glucose.