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A new class of hybrid biomaterials has recently evolved from the combination of peptides and DNA. In this chapter, we survey the unique systems and properties made possible by merging the bioactivity and mechanical properties of peptides with the programmability and reversibility of DNA. We explore peptide–DNA probes and switches, peptide-decorated DNA scaffolds and DNA-decorated peptide structures. We illustrate the rich variety of architectures made from the interplay of orthogonal peptide and DNA assembly motifs in cooperative assemblies, with examples of peptide-directed or DNA-directed hybrid structures. We present systems with hierarchical organization and dynamic assembly, presenting fascinating emergent properties made possible by peptide–DNA combinations. Finally, we discuss the future possibilities and open challenges presented by this intriguing class of hybrid biomaterials.

The single stranded DNA oligonucleotides known as aptamers have the capacity to bind proteins and other molecules and offer great therapeutic potential. Further work is required to optimize their function and to diminish their susceptibility to nuclease degradation. We report here on the synthesis and supramolecular self-assembly of DNA-peptide amphiphiles that form high aspect ratio nanofibers and display aptamers for platelet-derived growth factor. The nanofibers were found to bind the growth factor with an affinity that was fivefold greater than the free aptamer. We also observed that the aptamer displayed by the supramolecular nanostructures was eight times more nuclease resistant than free aptamer. In order to highlight the therapeutic potential of these supramolecular systems, we demonstrated the improved inhibition of proliferation when the growth factor was bound to aptamers displayed by the nanofibers.

DNA and peptides are two of the most commonly used biomolecules for building self-assembling materials, but few examples exist of hybrid nanostructures that contain both components. Here we report the modification of two peptides that comprise a coiled-coil heterodimer pair with unique DNA handles in order to link DNA origami nanostructures bearing complementary strands into micrometer-long one-dimensional arrays. We probed the effect of number of coils on self-assembly and demonstrated the formation of structures through multiple routes: one-pot assembly, formation of dimers and trimers and an alternating copolymer of two different origami structures, and stepwise assembly of purified structures with coiled-coil conjugates. Our results demonstrate the successful merging of two distinct self-assembly modes to create hybrid bionanomaterials expected to have a range of potential applications in the future.

The native extracellular matrix is a space in which signals can be displayed dynamically and reversibly, positioned with nanoscale precision, and combined synergistically to control cell function. Here we describe a molecular system that can be programmed to control these three characteristics. In this approach we immobilize peptide-DNA (P-DNA) molecules on a surface through complementary DNA tethers directing cells to adhere and spread reversibly over multiple cycles. The DNA can also serve as a molecular ruler to control the distance-dependent synergy between two peptides. Finally, we use two orthogonal DNA handles to regulate two different bioactive signals, with the ability to independently up- or downregulate each over time. This enabled us to discover that neural stem cells, derived from the murine spinal cord and organized as neurospheres, can be triggered to migrate out in response to an exogenous signal but then regroup into a neurosphere as the signal is removed., The extracellular matrix can affect cell behaviour both physically and biochemically. Here, the authors developed a substrate that is based on peptides and nucleic acids hybrids that can dynamically present signals upon demand which regulate cell adhesion and migration, thereby controlling cell organisation.

DNA and peptides are two of the most commonly used biomolecules for building self-assembling materials, but few examples exist of hybrid nanostructures that contain both components. Here we report the modification of two peptides that comprise a coiled-coil heterodimer pair with orthogonal DNA handles in order to link DNA origami nanostructures bearing complementary strands into micrometer long one-dimensional arrays. We probed the effect of number of coils on self-assembly and demonstrated the formation of self-assembled structures through multiple routes, to form dimers and trimers, an alternating copolymer of two different origami bundles, and stepwise assembly of purified structures with coiled-coil conjugates. Our results demonstrate the successful merging of two distinct self-assembly modes to create hybrid bionanomaterials expected to have a range of potential applications in the future.

Creating soft materials with the tunable hierarchical structures observed in nature remains an enormous challenge. Synthetic hierarchical systems have been reported, yet strategies to reversibly modulate their structure and function are scarce. We report on the programmable self-assembly of peptide-DNA brush copolymers into supramolecular architectures that can be tuned with changes in temperature, pH, or addition of a soluble trigger. A fiber to bundle transition occurs upon mixing peptides bearing complementary oligonucleotides. These hierarchical structures can be reversed using the programmable nature of DNA-DNA interactions. The ability to encode the final assemblies in the composition of both amino acid and DNA building blocks provides a strategy for constructing a unique class of reconfigurable supramolecular materials.

In this chapter, we will outline key applications for DNA as a scaffold for interfacing with and guiding the activity of biological systems. DNA is a highly programmable molecular building block but generally lacks activity for signaling cells. However, by synthesizing hybrid molecules of DNA linked to biologically active molecules like proteins or peptides, it is possible to imbue them with bioactivity. We will focus on two main categories of DNA-based materials: (1) three-dimensional hydrogels for cell encapsulation and/or delivery and (2) two-dimensional surfaces for studying cell-matrix interactions. Critically, we will restrict ourselves to systems that incorporate cells and strive to mimic the extracellular environment. These materials are highly promising for a broad range of applications in biology, bioengineering, and medicine.

We report the construction of DNA nanotubes covalently functionalized with the cell adhesion peptide RGDS as a bioactive substrate for neural stem cell differentiation. Alteration of the Watson–Crick base pairing program that builds the nanostructures allowed us to probe independently the effect of nanotube architecture and peptide bioactivity on stem cell differentiation. We found that both factors instruct synergistically the preferential differentiation of the cells into neurons rather than astrocytes.

This study describes the novel hemin/G-quadruplex DNAzyme-catalyzed aerobic oxidation of thiols to disulfides and the respective mechanism. The mechanism of the reaction involves the DNAzyme-catalyzed oxidation of thiols to disulfides and the thiol-mediated autocatalytic generation of H2O2 from oxygen. The coupling of a concomitant H2O2-mediated hemin/G-quadruplex-catalyzed oxidation of Amplex Red to the fluorescent resorufin as a transduction module provides a fluorescent signal for probing the catalyzed oxidation of the thiol to disulfides and for probing sensing processes that yield the hemin/G-quadruplex as a functional label. Accordingly, a versatile sensing method for analyzing thiols (L-cysteine, glutathione) using the H2O2-mediated DNAzyme-catalyzed oxidation of Amplex Red to the resorufin was developed. Also, the L-cysteine and Amplex Red system was implemented as an auxiliary fluorescent transduction module for probing recognition events that form the catalytic hemin/G-quadruplex structures. This is exemplified with the development of thrombin aptasensor. The thrombin/thrombin binding aptamer recognition complex binds hemin, and the resulting catalytic complex activates the auxiliary transduction module, involving the aerobic oxidation of l-cysteine and the concomitant formation of the fluorescent resorufin. Finally, the hemin/G-quadruplex DNAzyme/Amplex Red system was used to follow the activity of acetylcholine esterase, AChE, and to probe its inhibition. The AChE-catalyzed hydrolysis of acetylthiocholine to the thiol-functionalized thiocholine enabled the probing of the enzymatic activity of AChE through the hemin/G-quadruplex-catalyzed aerobic oxidation of thiocholine to the respective disulfide and the concomitant generation of the fluorescent resorufin product.

Graphene oxide (GO) is deposited onto the Al2O3 gates of field-effect transistor devices to yield a functional matrix for the sensing of thrombin or Hg2+ ions. The deposition of an aptamer subunit against thrombin or a tailored nucleic acid with appropriate T mismatches on the GO, yield active surfaces for the analysis of thrombin or Hg2+ ions. The desorption of the aptamer-subunit from GO, through the formation of the aptamer-thrombin complex, or the desorption of the T–Hg2+–T-bridged duplex DNA alter the gate potential, thus providing a signal for the sensing events.

A hemin/G-quadruplex nanostructure that is immobilized on CdS quantum dots (QDs) associated with an electrode leads, in the presence of luminol, H2O2, and triethanolamine as an electron donor, to the generation of photocurrents with no external irradiation of the QDs. The hemin/G-quadruplex-catalyzed generation of chemiluminescence leads to the chemiluminescence resonance energy transfer (CRET) to the QDs, resulting in the photoexcitation of the QDs and the generation of electron–hole pairs. The transfer of the conduction-band electrons to the electrode, and the concomitant scavenging of the valence-band holes by the triethanolamine electron donor result in the generation of photocurrents. The CRET-stimulated generation of photocurrents is applied to sense DNA by the labeling of the probe–analyte complex with a hemin/G-quadruple, and is also implemented to follow the activity of glucose oxidase and to sense glucose, by the labeling of the enzyme with the hemin/G-quadruplex catalyst.

The zinc(II)-protoporphyrin IX (ZnPPIX) fluorophore binds to G-quadruplexes, and this results in the enhanced fluorescence of the fluorophore. This property enabled the development of DNA sensors, aptasensors, and a sensor following telomerase activity. The DNA sensor is based on the design of a hairpin structure that includes a "caged" inactive G-quadruplex sequence. Upon opening the hairpin by the analyte DNA, the resulting fluorescence of the ZnPPIX/G-quadruplex provides the readout signal for the sensing event (detection limit 5 nM). Addition of Exonuclease III to the system allows the recycling of the analyte and its amplified analysis (detection limit, 200 pM). The association of the ZnPPIX to G-quadruplex aptamer-substrate complexes allowed the detection of adenosine-5'-triphosphate (ATP, detection limit 10 μM). Finally, the association of ZnPPIX to the G-quadruplex repeat units of telomers allowed the detection of telomerase activity originating from 380 ± 20 cancer 293T cell extract.

Graphene oxide (GO) is implemented as a functional matrix for developing fluorescent sensors for the amplified multiplexed detection of DNA, aptamer-substrate complexes, and for the integration of predesigned DNA constructs that activate logic gate operations. Fluorophore-labeled DNA strands acting as probes for two different DNA targets are adsorbed onto GO, leading to the quenching of the luminescence of the fluorophores. Desorption of the probes from the GO, through hybridization with the target DNAs, leads to the fluorescence of the respective label. By coupling exonuclease III, Exo III, to the system, the recycling of the target DNAs is demonstrated, and this leads to the amplified detection of the DNA targets (detection limit 5 × 10(-12) M). Similarly, adsorption of fluorophore-functionalized aptamers against thrombin or ATP onto the GO leads to the desorption of the aptamer-substrate complexes from GO and to the triggering of the luminescence corresponding to the respective fluorophore, thus, allowing the multiplexed analysis of the aptamer-substrate complexes. By designing functional fluorophore-labeled DNA constructs and their interaction with GO, in the presence (or absence) of nucleic acids, or two different substrates for aptamers, as inputs, the activation of the "OR" and "AND" logic gates is demonstrated.

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.

The ability of DNA to self-assemble into one-, two- and three-dimensional nanostructures, combined with the precision that is now possible when positioning nanoparticles or proteins on DNA scaffolds, provide a promising approach for the self-organization of composite nanostructures. Predicting and controlling the functions that emerge in self-organized biomolecular nanostructures is a major challenge in systems biology, and although a number of innovative examples have been reported, the emergent properties of systems in which enzymes are coupled together have not been fully explored. Here, we report the self-assembly of a DNA scaffold made of DNA strips that include 'hinges' to which biomolecules can be tethered. We attach either two enzymes or a cofactor-enzyme pair to the scaffold, and show that enzyme cascades or cofactor-mediated biocatalysis can proceed effectively; similar processes are not observed in diffusion-controlled homogeneous mixtures of the same components. Furthermore, because the relative position of the two enzymes or the cofactor-enzyme pair is determined by the topology of the DNA scaffold, it is possible to control the reactivity of the system through the design of the individual DNA strips. This method could lead to the self-organization of complex multi-enzyme cascades.

Phenyl boronic acid-functionalized CdSe/ZnS quantum dots (QDs) were synthesized. The modified particles bind nicotinamide adenine dinucleotide (NAD(+)) or 1,4-dihydronicotinamide adenine dinucleotide (NADH). The NAD(+)-functionalized QDs are effectively quenched by an electron transfer process, while the NADH-modified QDs are inefficiently quenched by the reduced cofactor. These properties enable the implementation of the QDs for the fluorescence analysis of ethanol in the presence of alcohol dehydrogenase. The NADH-functionalized QDs were used for the optical analysis of the 1,3,5-trinitrotriazine, RDX explosive, with a detection limit that corresponded to 1 x 10(-10) M. We demonstrate cooperative optical and catalytic functions of the core-shell components of the QDs in the analysis of RDX.

Dopamine (1) and tyrosinase (TR) activities were analyzed by using chemically modified ion-sensitive field-effect transistor (ISFET) devices. In one configuration, a phenylboronic acid functionalized ISFET was used to analyze 1 or TR. The formation of the boronate-1 complex on the surface of the gate altered the electrical potential associated with the gate, and thus enabled 1 to be analyzed with a detection limit of 7x10(-5) M. Similarly, the TR-induced formation of 1, and its association with the boronic acid ligand allowed a quantitative assay of TR to be performed. In another configuration, the surface of the ISFET gate was modified with tyramine or 1 to form functional surfaces for analyzing TR activities. The TR-induced oxidation of the tyramine- or 1-functionalized ISFETs resulted in the formation of the redox-active dopaquinone units. The control of the gate potential by the redox-active dopaquinone units allowed a quantitative assay of TR to be performed. The dopaquinone-functionalized ISFETs could be regenerated to give the 1-modified sensing devices by treatment with ascorbic acid.

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.

Appropriately ordered guanosine bases in DNA chains form G-quartets stabilized by interbase hydrogen bonds. These Gquartets are further stacked, in the presence of ions, into a Gquadruplex structure. Different organic molecules are known to interact with G-quadruplex structures. Specifically, hemin was found to bind to a G-quadruplex, and the resulting complex revealed horseradish peroxidise mimicking activities. For example, the hemin/G-quadruplex catalyzes the H2O2-mediated oxidation of the 2,2’-azino-bis(3-ethylbenzthiazoline-6-sulfonate dianion) (ABTS ) to the colored product ABTSC [3] or leads to the generation of chemiluminescence in the presence of H2O2/luminol. Also, the hemin/ G-quadruplex structure linked to electrodes was reported to act as an electrocatalyst for the reduction of H2O2, [5] and the hemin/G-quadruplex associated with semiconductor quantum dots was found to quench the luminescence of the quantum dots. These properties enabled the use of the hemin/Gquadruplex as a versatile label for numerous sensors including enzyme-, DNA-, and aptamer-based sensors. In the present study, we demonstrate that the hemin/G-quadruplex acts not only as a horseradish peroxidase mimicking DNAzyme, but, also, as an NADH oxidase and NADH peroxidase mimicking DNAzyme. The regeneration of the nicotinamide adenine dinucleotide, NAD, cofactor attracted substantial interest for biotechnological applications, and NAD-dependent enzymes have been widely used for chemical transformations. Our results pave the way to use the hemin/Gquadruplex as a biocatalyst for the regeneration of the NAD cofactor and to apply the DNAzyme as a catalyst for enzymedriven transformations. NADH oxidase catalyzes the oxidation of NADH by O2 with the concomitant formation of H2O2. [11] The present system consists of reduced nicotinamide adenine dinucleotide (NADH), hemin/G-quadruplex (1), and Amplex Red (2), as a fluorescent reporter dye (Scheme 1A). Under aerobic conditions, NADH is oxidized to NAD (see below), while 2 is oxidized to resorufin (3), which generates fluorescence at lmax= 581 nm. Figure 1A, curve (a), shows the time-dependent increase in the fluorescence generated by the system. Control experiments reveal minor background fluorescence generated under an inert argon atmosphere (curve (e)), or upon exclusion of hemin from the system (curve (f)). Also, only a low fluorescence intensity is generated in the presence of only hemin and in the absence of 1 (curve (d)), or in the presence of hemin and a foreign nucleic acid (4) that cannot form the respective G-quadruplex (curve (c)). These results may suggest that the hemin/G-quadruplex-catalyzed oxidation of NADH by O2 yields H2O2, and the generated fluorescence occurs by the well-established hemin/G-quadruplex DNAzyme-catalyzed oxidation of 2 by H2O2. [12] Indeed, the intermediate formation of H2O2 in the system was confirmed by the addition of catalase. Figure 1A, curve (g), confirms that in the presence of catalase the fluorescence is blocked, consistent with the decomposition of the generated H2O2. Thus, in the first step, the hemin/Gquadruplex DNAzyme acts as an NADH oxidase, where NADH is oxidized by O2 to form NAD . It should be noted that the fluorescent resorufin (3) is formed also in the absence of NADH (Figure 1A, curve (b)). Presumably, Amplex Red by itself has a residual donor activity that participates in the activation of O2 (substituting NADH). The low fluorescence intensity changes should be considered as the background signal of the system. Scheme 1. A) Frame I: Hemin/G-quadruplex-catalyzed oxidation of NADH by O2 to NAD + and H2O2, respectively, and the concomitant oxidation of Amplex Red (2) to resorufin (3) by H2O2. Frame II: Coupled DNAzyme–alcohol dehydrogenase biocatalytic system for the formation of NADH and the concomitant regeneration of the NAD cofactor by the hemin/G-quadruplex. B) Suggested mechanism for the DNAzyme-catalyzed oxidation of NADH.

The incorporation of hemin into the thrombin/G-quadruplex aptamer assembly or into the ATP/G-quadruplex nanostructure yields active DNAzymes that catalyze the generation of chemiluminescence. These catalytic processes enable the detection of thrombin and ATP with detection limits corresponding to 200 pM and 10 μM, respectively. The conjugation of the antithrombin or anti-ATP aptamers to CdSe/ZnS semiconductor quantum dots (QDs) allowed the detection of thrombin or ATP through the luminescence of the QDs that is powered by a chemiluminescence resonance energy-transfer (CRET) process stimulated by the hemin/G-quadruplex/thrombin complex or the hemin/G-quadruplex/ATP nanostructure, in the presence of luminol/H(2)O(2). The advantages of applying the CRET process for the detection of thrombin or ATP, by the resulting hemin/G-quadruplex DNAzyme structures, are reflected by low background signals and the possibility to develop multiplexed aptasensor assays using different sized QDs.

The optical detection of DNA or the sensing of low-molecular-weight substrates or proteins by aptamer nucleic acids is a long term challenge in the design of biosensors. Similarly, the detection of the telomerase activity, a versatile biomarker of cancer cells, is important for rapid cancer diagnostics. We implement the luminescence quenching of the CdSe/ZnS quantum dots (QDs) as a versatile process to develop DNA sensors and aptasensors, and to design an analytical platform for the detection of telomerase activity. The formation of nucleic acid duplexes on QDs, or the assembly of aptamer-substrate complexes on the QDs (substrate=cocaine or thrombin) is accompanied by the intercalation of doxorubicin (DB) into the duplex domains of the resulting recognition complexes. The intercalated DB quenches the luminescence of the QDs, thus leading to the detection readout signal. Similarly, the telomerase-induced formation of the telomere chains on the QDs is followed by the hybridization of nucleic-acid units complementary to the telomere repeat units, and the intercalation of DB into the resulting duplex structure. The resulting luminescence quenching of the QDs provides an indicating signal for the activity of telomerase.

The luminescence of CdSe/ZnS QDs is quenched via electron transfer by hemin/G-quadruplex associated with the particles. This phenomenon is implemented to develop DNA sensors or aptasensors by tailoring hairpin-functionalized QDs that generate the hemin/G-quadruplex quenchers upon sensing of the respective analytes.

This study demonstrates the self-assembly of inhibitor/enzyme-tethered nucleic acid fragments or enzyme I-, enzyme II-modified nucleic acids into functional nanostructures that lead to the controlled inhibition of the enzyme or the activation of an enzyme cascade. In one system, the anti-cocaine aptamer subunits are modified with monocarboxy methylene blue (MB(+)) as the inhibitor and with choline oxidase (ChOx). The cocaine-induced self-assembly of the aptamer subunits complex results in the inhibition of ChOx by MB(+). In a further configuration, two nucleic acids of limited complementarity are functionalized at their 3' and 5' ends with glucose oxidase (GOx) and horseradish peroxidase (HRP), respectively, or with MB(+) and ChOx. In the presence of a target DNA sequence, synergistic complementary base-pairing occurs, thus leading to stable supramolecular Y-shaped nanostructures of the nucleic acid units. A GOx/HRP bienzyme cascade or the programmed inhibition of ChOx by MB(+) is demonstrated in the resulting nucleic acid nanostructures. A quantitative theoretical model that describes the nucleic acid assemblies and that results in the inhibition of ChOx by MB(+) or in the activation of the GOx/HRP cascade, respectively, is provided.

Beta-cyclodextrin (beta-CD)-functionalized CdSe/ZnS quantum dots (QDs) are used for optical sensing and chiroselective sensing of different substrates using a fluorescence resonance energy transfer (FRET) or an electron transfer (ET) mechanisms. The FRET between the QDs and Rhodamine B incorporated in the beta-CD receptor sites is used for the competitive analysis of adamantanecarboxylic acid and of p-hydroxytoluene. Also, the dye-incorporated beta-CD-modified QDs are used for the chiroselective optical discrimination between D,L-phenylalanine and D,L-tyrosine. The receptor-functionalized QDs are also implemented for the optical detection of p-nitrophenol using an ET quenching route.

The fluorescence resonance energy transfer (FRET) between CdSe-ZnS QDs and fluorophore-labeled galactose or fluorophore-labeled dopamine linked to phenyl boronic acid-functionalized QDs is used to develop competitive assays for the optical detection of galactose, glucose, or dopamine.

The electrochemical and photoelectrochemical detection of tyrosinase (TR) activity (an indicative marker for melanoma cancer cells) is reported, using Pt nanoparticles (NPs) or CdS NPs as electrocatalytic labels or photoelectrochemical reporter units. The Pt NPs or CdS NPs are modified with a tyrosine methyl ester, (1), capping layer. Oxidation of the capping layer by TR/O2 yields the respective L-DOPA and dopaquinone products. The reduction of the resulting mixture of products with citric acid yields the L-DOPA derivative,(3), as a single product. The association of the (3)-functionalized Pt NPs or CdS NPs to a boronic acid monolayer-modified electrode enables the electrochemical transduction of TR activity by the Pt-NPs-electrocatalyzed reduction of H2O2 or the photoelectrochemical transduction of TR activity by the generation of photocurrents in the presence of triethanolamine as a sacrificial electron donor. The detection limits for analyzing TR corresponds to 1 U and 0.1 U by the electrochemical and photoelectrochemical methods, respectively. The association of the Pt NPs or CdS NPs to the functionalized monolayer electrode is followed by quartz crystal microbalance measurements.

Hemin/G-quadruplex catalyzes the H(2)O(2)-mediated oxidation of Amplex Red to the fluorescent product resorufin. This process is implemented to develop hairpin nucleic acid structures for the detection of DNA, to probe the catalytic activity of glucose oxidase, to use the thrombin-aptamer complex as a catalytic readout structure, and to quantitatively analyze telomere chain composition.

The anti-cocaine aptamer was fragmented into two nucleic acids, (1) and (2). The nucleic acid (1) was tethered at its 5'-end to aminoethyl nicotinamide adenine dinucleotide, amino-NAD(+), or to horseradish peroxidase, HRP. The nucleic acid (2) was functionalized at its 3'-end with alcohol dehydrogenase, AlcDH, or with glucose oxidase, GOx. In the presence of cocaine, the supramolecular NAD(+)/AlcDH/cocaine-aptamer complex is formed, and the biocatalytic oxidation of ethanol is activated. Similarly, in the presence of cocaine, the GOx/HRP/cocaine-aptamer complex is formed, and this activates the biocatalytic cascade where glucose is oxidized by GOx to yield gluconic acid and H(2)O(2), and the resulting hydrogen peroxide activates the HRP-biocatalyzed oxidation of 2,2'-azinobis-(3-ethylbenzthiazoline-6-sulfonate), ABTS(2-). The systems may be considered as biomimetic prototypes for systems biology.

The activities of the enzymes tyrosinase and thrombin are probed by the association of the ferrocene boronic acid label to the enzyme-generated catechol ligand, and by the cleavage of the ligand-redox complex tethered to a peptide, respectively.