DNA and telomerase activity are detected by a DNAzyme generated upon hybridization and opening of a functional catalytic beacon.

DNAzyme-functionalized Au−NPs act as catalytic labels for the amplified detection of DNA and telomerase activity on nucleic acid-functionalized gold surface. The DNAzyme stimulates, in the presence of hemin, H 2 O 2 , and luminol, the generation of chemiluminescence. For DNA analysis, a nucleic acid unit complementary to the analyzed DNA is tethered to the DNAzyme structure associated with the Au−NPs. For telomerase activity, a nucleic acid complementary to the telomer repeat units generated on the surface is tethered to the DNAzyme structure associated with the Au−NPs. The detection limit for the detection of DNA is 1 × 10 -10 M. The method enables the detection of telomerase activity originating from 1000 HeLa cells.

Different selected enzymes, glucose oxidase (GOx), catalase (Cat), glucose dehydrogenase (GDH), horseradish peroxidase (HRP), and formaldehyde dehydrogenase (FDH), are used alone or coupled to construct eight different logic gates. The added substrates for the respective enzymes, glucose and H(2)O(2), act as the gate inputs, while the biocatalytically generated gluconic acid or NADH are the output signals that follow the operation of the gates. Different enzyme-based gates are XOR, INHIBIT A, INHIBIT B, AND, OR, NOR, Identity and Inverter gates. By combining the AND and XOR or the XOR and INHIBIT A gates, the half-adder and half-subtractor are constructed, respectively, opening the way to elementary computing by the use of enzymes.

The assembly of three concatenated enzyme-based logic gates consisting of OR, AND, XOR is described. Four biocatalysts, acetylcholine esterase, choline oxidase, microperoxidase-11, and the NAD ؉ -dependent glucose dehydrogenase, are used to assemble the gates. Four inputs that include acetylcholine, butyrylcholine, O2, and glucose are used to drive the concatenated-gates system. The cofactor NAD ؉ , and its reduced 1,4-dihydro form, NADH, are used as a reporter couple, and these provide an optical output for the gates. The modulus of the absorbance changes of NADH is used as a readout signal.biocomputers ͉ biocatalysis ͉ enzymes T he hardware of computers consists of parallel and serial logic-gate operations that are triggered by electronic inputs. These functions may be duplicated by appropriately designed chemical or biological systems. Different molecular and supramolecular assemblies that operate as logic gates and perform molecular-scale arithmetic operations were discussed (1-9). Similarly, biomolecules such as nucleic acids or proteins were used as active components that perform logic-gate operations (10-15). Gene-based artificial circuits acting as bistable toggle switches (16) or oscillators (17) were developed, and coupled enzyme͞DNA systems that perform programmable biochemical transformations that mimic basic computing of finite automaton were reported (17). The use of enzymes as the active components for logic gate functions is specifically intriguing because numerous biocatalytic cycles in nature rely on information processing, revealing similarities to computer devices. Although the function of enzyme networks as mimics of Boolean logic gates was discussed (18), and the potential use of enzymes as building units of high-density computing architectures was addressed (19), the experimental work that validates biocatalyst-stimulated logic gate operations is quite scarce, and lacks the desired complexity that resembles computers. Single enzyme-based logic-gate operations were reported. For example, the dynamic conformational changes of malate dehydrogenase in response to Mg ϩ and Ca 2ϩ ions acting as inputs was used to develop a XOR gate (20). Also, a modified enzyme and its inhibitor were used as inputs that activated an AND gate (21). Recently, we have reported on the assembly of coupled biocatalytic systems that mimic OR, XOR, AND, or InhibAND logic-gate functions (22), and the use of these systems for elementary arithmetic operations (halfadder and half-subtractor) was demonstrated (23). In none of these systems was the consecutive operation of several gates that operate in series demonstrated. This feature is, however, essential to develop any future ''computer-like'' function of enhanced complexity. Here, we report on the assembly of a four-enzymecoupled system that includes four inputs and performs in series three logic-gate operations OR, AND, and XOR. Results and DiscussionsThe system and its operation is depicted in Fig. 1A and consists of the four biocatalysts, acetylcholine esterase (AChE)...

Physio‐logical: A system of four enzymes, horseradish peroxidase, glucose dehydrogenase, glucose oxidase, and catalase, operate in parallel in the presence of NADH and NAD+ to yield AND and XOR gates (see picture). Glucose and H2O2 act as the inputs, and the absorbance that follows the production of gluconic acid or NADH provide the outputs of the AND and XOR gates, respectively.

Increasing efforts are directed to the application of metal and semiconductor nanoparticles (NPs) for the development of electronic or optical sensory systems.[1] Metal or semiconductor NPs functionalized with nucleic acids were employed as amplifying labels for the detection of DNA; the dissolution of the nanoparticles was used to follow DNA hybridization events.[2] Also, charge injection from semiconductor nanoparticles into electrodes and the generation of photocurrents was used to follow hybridization processes [3] and biocatalytic transformations.[4] The catalytic deposition of metals onto metal nanoparticles conjugated to DNA-hybridized complexes on surfaces was used as a sensor for DNA through conductivity [5] or microgravimetric quartz crystal microbalance [6] measurements. The optical detection of processes in the presence of metal and semiconductor NPs has become a common practice in analysis. Besides the use of semiconductor quantum dots as fluorescence labels in sensors, [7] the fluorescence quenching of semiconductor quantum dots has been employed in different sensing paths.[8] The plasmon absorbance of metal nanoparticles, such as Au NPs, and specifically the interparticlecoupled plasmon absorbance of aggregated NPs was extensively used to follow molecular [9] and biomolecular [10] recognition processes. The use of semiconductor or metallic NPs as probes to follow biocatalytic processes is less established, with only a few reports for these applications. [4,11] Nicotinamide adenine dinucleotide (NAD + )-and nicotinamide adenine dinucleotide phosphate (NADP + )-dependent enzymes are important in biocatalyzed synthesis.[12] Extensive efforts have been directed towards the development of electrochemical sensors based on NAD(P) + -dependent enzymes.[13] Herein, we report the catalyzed growth of gold nanoparticles in the presence of NAD(P)H cofactors. We apply the process to the quantitative optical analysis of NAD(P)H cofactors and to the analysis of NAD(P) + -dependent biocatalyzed reactions in solutions and on surfaces.The solution for the growth of the particles consisted of citrate-stabilized Au NPs (4.0 10 À10 m in 13 nm AE 1-nm particles), HAuCl 4 (1.8 10 À4 m), and CTAB (7.4 10 À2 m) as a surfactant. Figure 1 shows the changes in the UV/Vis spectra of the growth solution upon interaction with different concentrations of NADH. In the absence of NADH, the solution displays an absorbance band at l = 392 nm, characteristic of the AuCl 4 À component (Figure 1, curve a). Upon addition of NADH, this band disappears instantaneously and the characteristic orange color of the system is depleted (curve b), and then the slow buildup of the absorbance of the particle plasmon is observed. As the concentration of NADH increases, the absorbance of the Au particles increases and is shifted to longer wavelengths (from 523 to 530 nm; Figure 1, curves c-h). The inset in Figure 1 shows the calibration curve derived from the changes in the absorbance at l = 524 nm as the concentration of NADH increases. Figure 2 shows...

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Physio‐logisch: Vier Enzyme – Meerrettichperoxidase, Glucose‐Dehydrogenase, Glucose‐Oxidase und Katalase – arbeiten in Gegenwart von NADH und NAD+ parallel in AND‐ und XOR‐Schaltungen (siehe Bild). Glucose und H2O2 dienen als Eingabe, und Absorbanzänderungen infolge der Produktion von Gluconsäure oder NADH werden als Ausgabe der AND‐ bzw. XOR‐Schaltung wahrgenommen.