functional dna nanotechnology: emerging applications of dnazymes and aptamers

Functional DNA nanotechnology: emerging applications ofDNAzymes and aptamersYi Lu and Juewen Liu

In the past 25 years, DNA molecules have been utilized both as

powerful synthetic building blocks to create nanoscale

architectures and as versatile programmable templates for

assembly of nanomaterials. In parallel, the functions of DNA

molecules have been expanded from pure genetic information

storage to catalytic functions like those of protein enzymes

(DNAzymes) and specific binding functions like antibodies

(aptamers). In the past few years, a new interdisciplinary field

has emerged that aims to combine functional DNA biology with

nanotechnology to generate more dynamic and controllable

DNA-based nanostructures or DNA-templated nanomaterials

that are responsive to chemical stimuli.

AddressesDepartment of Chemistry, Beckman Institute for Advanced Science

and Technology, University of Illinois at Urbana-Champaign, Urbana,

IL 61801, USA

Corresponding author: Lu, Yi ([email protected])

Current Opinion in Biotechnology 2006, 17:580–588

This review comes from a themed issue on

Chemical biotechnology

Edited by Jonathan S Dordick and Amihay Freeman

Available online 23th October 2006

0958-1669/$ – see front matter

# 2006 Elsevier Ltd. All rights reserved.

DOI 10.1016/j.copbio.2006.10.004

IntroductionTraditionally, DNA molecules were thought to have the

sole function of carrying and passing genetic information

from one generation to another. About 25 years ago, how-

ever, DNA began to find a new role in the field of materials

science [1–6]. Structurally, DNA can bind specifically to

another DNA strand of complementary sequence, with the

double-stranded DNA forming a solid rod of up to�50 nm

in length. Moreover, DNA can be modified with a wide

range of fluorophores and other functional groups that

enable it to conjugate with nanoparticles. Compared with

RNA and proteins, DNA molecules are less susceptible to

hydrolysis and thus are highly stable. These attributes

make DNA a special biopolymer with highly predictable

sequence-dependent properties, and these properties

have been exploited to construct DNA-based geometric

and topologic structures [4,7]. By using ‘sticky ends’,

dispersed three- or four-way DNA junctions and other

DNA building blocks have been connected into large

periodic structures, upon which nanoparticles or proteins


can be deposited to form well-defined patterns [8,9].

Alternatively, DNA has been conjugated to inorganic

nanoparticles, which can then be assembled in a program-

mable manner to form structures containing either a lim-

ited number of particles or cross-linked nanoparticle

aggregates [1,2,10,11]. These research efforts have demon-

strated the power of DNA as a structural molecule, a

scaffold, and as a template in developing nanotechnology.

Almost in parallel with the development of DNA nano-

technology, the chemical functions of DNA have been

expanded beyond the DNA double helix. Since the early

1990s, many DNA molecules — known as DNA aptamers

— have been isolated that are able to bind a broad range

of molecules with high affinity and specificity [12,13].

The molecules that can be recognized by aptamers range

from small organic molecules to proteins, cells and even

intact viral particles [14]. A further advance in the devel-

opment of function DNAs was made in 1994, when DNA

was shown to act as a catalyst for the first time [15].

Herein, these catalytic DNA molecules are called DNA-

zymes (also described as DNA enzymes, deoxyribozymes

or catalytic DNA elsewhere). Moreover, DNAzymes and

aptamers have been combined to form allosteric DNA-

zymes or aptazymes. These DNAzymes, aptamers and

aptazymes are collectively called functional DNAs,

whose functions extend beyond the Watson–Crick base

pair recognition of complementary strands [16�].

Given the tremendous progress made in both DNA

nanotechnology and in the study of functional DNA, it

is natural to combine these two exciting fields to create a

new interdisciplinary field that uses functional DNA to

control and fine-tune the structure and dynamics of DNA

nanostructures and materials. This is exactly what has

happened in the past few years [17]. Because new DNA

functions always involve interactions with other chemical

and biological molecules, they allow the generation of

‘smart’ nanoscale architectures; the assembly and func-

tion of these architectures are responsive to chemical or

biological stimuli, in much the same way that biomaterials

are made and function in cells. Compared with DNA

nanotechnology based on base-pair hybridizations, nano-

technologies based on functional DNA could potentially

be more versatile and dynamic. The change in chemical

or physical properties of the devices can be used to detect

those stimuli in a highly sensitive and selective manner.

The sensing applications of functional DNA have been

recently reviewed [17,18] and this review mainly focuses

on recent progress in other aspects of functional DNA

nanotechnology.

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http://dx.doi.org/10.1016/j.copbio.2006.10.004

Functional DNA nanotechnology Lu and Liu 581

DNAzymes in nanotechnologyGeneral considerations

DNAzymes are DNA-based biocatalysts capable of per-

forming chemical transformations [15,19–22]. These cat-

alysts have not been found in nature, and all known

DNAzymes were isolated by in vitro selection. So far,

most of their substrates have been found to be nucleic

acids. DNAzymes can therefore provide additional con-

trol over nucleic-acid-based nanodevices and, because

DNAzymes often catalyze multiple turnover reactions,

such devices can have amplification effects. Among the

many classes of DNAzymes, RNA-cleaving DNAzymes

Figure 1

DNAzyme-templated material assembly. (a) Secondary structure of the Pb2+

by the DNAzyme in the presence of Pb2+. Pb2+-directed assembly of gold n

a head-to-tail [23��] or (d) a tail-to-tail manner [25]. The nanoparticles are de

on assembly, owing to surface plasmon effects. (e) For head-to-tail aligned

change can be observed. (f) For tail-to-tail aligned aggregates, Pb2+ can ind

and stable DNAzyme–nanotube hybrid [29�]. The DNAzyme is immobilized o

reactions with high turnover numbers.


are the most widely used, mainly because of their simple

reaction conditions, fast turnover rates, and significant

modifications on their substrate lengths.

DNAzyme-templated material assembly

Liu and Lu [17,23��,24,25,26��,27��,28] were the first to

employ DNAzymes to obtain functional nanomaterials

that are sensitive to chemical stimuli. Their work

employed the use of a Pb2+-specific DNAzyme (shown

bound to its substrate DNA in Figure 1a). In the presence

of Pb2+, the DNAzyme (enzyme strand; green) cleaves

the substrate strand (black) into two pieces (Figure 1b). In

-specific DNAzyme discussed in the text. (b) Cleavage of the substrate

anoparticles by the DNAzyme when nanoparticles are aligned (c) in

picted in red or blue; a red to blue/purple color transition is observed

aggregates, Pb2+ cannot induce DNAzyme cleavage and no color

uce DNAzyme cleavage and color change [26��]. (g) A highly active

n a carbon nanotube and the hybrid can perform efficient cleavage


582 Chemical biotechnology

order to assemble nanoparticles, the substrate strand was

extended at both ends with segments that were comple-

mentary to DNA fragments attached to the nanoparticles.

On assembly of the nanoparticles into an aggregate, a red

to blue colour change is observed that results from a shift

in the surface plasmon resonance properties of the par-

ticle. If the nanoparticles were aligned in a head-to-tail

manner (Figure 1c), a heating and cooling (annealing)

step was needed to form blue-colored aggregates [23��].In the presence of Pb2+, the assembly was inhibited

because of substrate cleavage. Changing the alignment

to tail-to-tail (Figure 1d) allowed the assembly to occur at

ambient temperatures [25]. The reverse process — Pb2+-

induced disassembly of nanoparticle aggregates — was

also studied [26��,28]. Interestingly, if the particles were

aligned head-to-tail, the DNAzyme was not active

(Figure 1e). Activity and disassembly of nanoparticles

were observed for tail-to-tail aligned aggregates

(Figure 1f). Because of the accompanying red/blue color

transitions, these materials are useful for the colorimetric

sensing of Pb2+ or other metal ions [17,23��,24,25,26��,28].

These research efforts have led to the use of DNAzymes

to proofread and correct errors or defects in nanomaterial

assembly, an important task for practical applications of

nanomaterials [27��].

The same Pb2+-specific DNAzyme was also immobilized

onto carbon nanotubes in collaborative work between the

groups of Lu, Kane and Dordick (Figure 1g) [29�]. The

immobilized DNAzyme formed a highly stable hybrid

with nanotubes and maintained high activity; over 400

turnovers were observed for each molecule of DNAzyme.

Such high activity could facilitate many applications,

ranging from the directed assembly of nanotubes to

nanoscale cellular therapeutics. Along the same lines of

surface immobilization, the DNAzyme was covalently

attached to gold surfaces [30] and gold-coated nanocapil-

lary membranes. In this case, enzyme activity was main-

tained even after storage in a dried state for 30 days at

room temperature [31].

Micro- and nanofluidic devices are promising platforms

for several applications, including sensing and drug deliv-

ery. The use of DNAzymes in such devices would enable

the analysis of extremely small samples, unattended long-

term monitoring, and potential multiplex array sensing.

The Pb2+-specific DNAzyme discussed above was placed

in a microfluidic device [32] and less than 1 nL of DNA

was found to be needed to monitor the cleavage reaction

for Pb2+ sensing. The DNAzyme was also assayed in a

nanofluidic device containing two perpendicular channels

interfaced by a nanocapillary array interconnect. This

voltage-controlled device was capable of detecting Pb2+

concentrations as low as 11 nM [33].

Seeman, Ellington and co-workers [34] recently incorpo-

rated a Cu2+-dependent DNAzyme [35] with optimized


cleavage efficiency [34] into a two-dimensional array

based on the DNA double crossover motif (a rigid

DNA building block frequently used in DNA nanotech-

nology). The array was designed to contain two types of

alternating stripes: one from the DNAzyme and the other

from an embedded DNA hairpin. The separation

between the stripes was 32 nm. After addition of Cu2+,

the DNAzyme was cleaved off the array and the separa-

tion changed to 64 nm [34]. The authors propose that

many DNAzymes responsive to different chemical sti-

muli could be used to construct such nanostructures the

properties of which can then be controlled by those

stimuli in a programmable manner.

DNAzyme-based DNA molecular devices

Mao and co-workers [36,37�,38��] have used DNAzymes

to construct molecular motors with open-close motions

and walking motions. For example, a DNAzyme

(Figure 2a, in green) was flanked by two double-stranded

overhangs to form a closed structure. The addition of

substrate DNA (in purple) forced the device to open.

Subsequently, the substrate was cleaved by the DNA-

zyme into two pieces and released, which moved the

device back to the closed state. The device will continue

to operate as long as free substrate is present [36]. By

introducing a non-cleavable substrate, the motion of the

device can be stopped; removal of the non-cleavable

substrate can resume the motion [37�].

A DNAzyme was also designed to walk along a DNA

track (Figure 2b) [38��]. Initially, the DNAzyme was

hybridized to the substrate DNA, but the hybridization

was designed to be asymmetric with one arm being longer

than the other; one end of the substrate was hybridized to

a DNA track (in blue). Cleavage of the substrate released

the shorter substrate fragment leaving the corresponding

free DNAzyme portion available to seek a nearby free,

full-length substrate. Because the stability of hybridiza-

tion to the full-length substrate was higher, the DNA-

zyme moved completely to a new substrate to perform a

second cleavage reaction. Up to three migration steps

have been demonstrated with the device.

Aptamers in nanotechnologyGeneral considerations

In parallel to protein-based antibodies, aptamers are

nucleic-acid-based binding molecules [12–14]. Aptamers

are obtained by a combinatorial selection method known

as systematic evolution of ligands by exponential enrich-

ment (SELEX), in which DNA molecules with the

desired binding properties are isolated from a library

containing as many as 1015 random sequences. What

makes aptamers unique and useful in nanotechnology

is the fact that an aptamer can be controlled in three

distinct states: free in solution, bound to its target mole-

cule or bound to its complementary DNA [39,40��].Switching between the three states can generate



Figure 2

DNAzyme-based devices. (a) A nanomotor with open-close motors powered by a DNAzyme [36,37�]. When the DNAzyme (in green) is free of

substrate, the device is in the closed state. Hybridization to a substrate (in purple) opens the device, which is subsequently closed by DNAzyme

cleavage and product release. (b) A DNAzyme nanowalker [38��]. A DNAzyme (in green) is initially hybridized to the first substrate (in purple)

immobilized on a DNA track (blue). Cleavage of the substrate induces migration of the DNAzyme to a nearby new substrate.

mechanical motions, induce or release constraints, and

associate or dissociate chemicals or oligonucleotides.

Many recent studies have taken advantage of this feature.

Aptamer-templated material assembly

Aptamers have been recently employed to assemble

nanoparticles, and many of the resulting materials are

dynamic in nature and sensitive to chemical stimuli. In

one such study, Willner and co-workers [41] functiona-

lized gold nanoparticles with an antithrombin aptamer.

In the presence of high concentrations of thrombin, the

nanoparticle solution showed some turbidity, which was

attributed to each thrombin molecule interacting with

two thrombin aptamers [42] and initiating nanoparticle

assembly to form aggregates (Figure 3a). Similarly,

Chang and co-workers employed platelet-derived growth

factor (PDGF) to assemble PDGF-aptamer-functiona-

lized gold nanoparticles with a visible red-to-purple color

change in the presence of PDGF [43�]. As in the DNA-

zyme studies described earlier, the color change results

from changes in the surface plasmon resonance coupling

of the nanoparticles on binding. These assemblies are

limited to molecules that can bind multiple aptamers. In

a complementary study, Liu and Lu [44��] reported a

more general design in which aptamers were used as a

linker to assemble DNA-functionalized gold nanoparti-

cles (Figure 3b). In the presence of target molecules, the

aptamer switched its structure and the nanoparticles

dissociated. As a result, purple-colored aggregates dis-

persed into red colored individual nanoparticles. This

design can be used to obtain nanomaterials sensitive to a

broad range of molecules by simple replacement of the

aptamer sequences. Nanostructures sensitive to adeno-

sine, cocaine and potassium ions have been demon-


strated [44��,45��]. Because of its generality, more

complex systems were built to be responsive to multiple

chemical inputs with controllable input cooperativity

[45��]. For example, the disassembly of materials shown

in Figure 3c required the presence of both adenosine and

cocaine, whereas the disassembly of materials shown in

Figure 3d can happen with either molecule. Besides

nanoparticle aggregates, protein arrays have also been

made by aptamer-directed assembly. For instance, Yan

and co-workers [46�] used thrombin aptamers to form

one-dimensional thrombin arrays (Figure 3e). Because

aptamers are also a fragment of DNA, such a design can

be conveniently incorporated into the construction of a

DNA scaffold without the need to introduce other func-

tional groups for protein conjugation. Incorporation of a

fluorophore-modified aptamer into such DNA arrays

enabled fluorescent detection of thrombin (see also

Update).

Aptamers have also been applied to control the chemical

and physical properties of other materials. Ellington and

co-workers [47�] employed aptamers to control the emis-

sion of semiconductor nanocrystals called quantum dots

(QDs) (Figure 3f). A short piece of quencher-labeled

DNA was hybridized to a thrombin aptamer attached

to a QD; therefore, the QD emission was initially

quenched. Addition of thrombin resulted in dissociation

of the quencher-labeled DNA and unmasked the QD

emission. Willner and co-workers [48] also made use of

thrombin aptamers — in this case for the immobilization

of platinum (Pt) nanoparticles. The authors immobilized

thrombin aptamers on a gold surface and functionalized

Pt nanoparticles with thrombin aptamers (Figure 3g) [48].

Thrombin was then sandwiched between the two



Figure 3

Aptamer-templated nanomaterial assembly. (a) The assembly of thrombin-aptamer-functionalized gold nanoparticles by thrombin [41]. Each thrombin

molecule (yellow) can bind two aptamers. The same concept has also been used to assemble platelet-derived growth factor (PDGF)-aptamer-

functionalized gold nanoparticles [43�]. The nanoparticles are depicted in red or blue; a red to blue/purple color transition is observed on assembly,

owing to surface plasmon effects. (b) Aptamer-assembled nanoparticle aggregates sensitive to chemical inputs such as adenosine, cocaine

and K+ [44��]. Upon binding to adenosine, the nanoparticles are dissociated into the dispersed state. Nanomaterials responsive to adenosine

and cocaine with (c) high or (d) no cooperativity [45��]. (e) Patterning thrombin arrays using thrombin aptamers as a capturing agent [46�].

(f) Aptamer-controlled emission properties of quantum dots (QDs) [47�]. A quencher-labeled DNA is positioned close to the QD by hybridization

to an attached thrombin aptamer. Addition of thrombin releases the quencher labeled DNA, unmasking the fluorescence. (g) Thrombin-mediated

immobilization of Pt nanoparticles on a gold surface for catalyzing the reduction of H2O2 [48].

aptamers and, as a result, the Pt nanoparticle was immo-

bilized. Pt nanoparticles can catalyze the reduction of

H2O2 to H2O to generate electrochemical signals. This

system is useful for the amplified electrochemical detec-

tion of thrombin.

Aptamer-based DNA molecular devices

Most DNA-based molecular devices are powered by the

energy generated from DNA hybridization. By contrast,

in a biological system, most motors are fueled by ATP

hydrolysis. Recent work on aptamer-based devices has

shown great promise in mimicking biological systems.

The first example, which can be considered an aptamer-

based DNA motor, was reported by Li and Tan [49]. A

piece of guanine-rich DNA (Figure 4a, top in green) can


form a quadruplex by binding to K+, and this piece of

DNA can be considered a K+ aptamer. The addition of

complementary DNA (in blue) forces the quadruplex to

open leaving the DNA in a duplex form. By leaving a tail

on the added DNA, when the full complementary DNA

(in orange) is added, the K+ aptamer DNA is released to

re-associate with K+ and form a quadruplex structure. The

aptamer can be switched in the state of binding to K+ or

binding to its complementary DNA for over ten cycles.

The same sequence can also bind thrombin [50]. Simmel

and co-workers [51�] have extended the quadruplex

region with a tail (Figure 4b, in black) to facilitate the

release of thrombin by a complementary strand. As in the

earlier study, the aptamer DNA switched between bind-

ing to thrombin and to the complementary DNA for



Figure 4

Aptamer-based DNA devices. (a) A DNA nanomotor powered by K+ and DNA [49]. The motor (in green) adopts a quadruplex structure by binding

K+ (red). In the presence of complementary DNA (blue), the motor becomes part of a DNA duplex. Cycling of the motor is realized by the addition

of another DNA (orange) to free the motor. (b) A DNA nanomotor powered by thrombin and DNA [51�]. The motor (in green with a black tail)

adopts a quadruplex structure by binding thrombin. In the presence of complementary DNA, the motor becomes part of a DNA duplex and leaves

the thrombin. Cycling of the motor is realized by the addition of another DNA (in orange) to free the motor. (c) A DNA device powered by adenosine

and adenosine deaminase [52��]. A short DNA molecule (in blue) is hybridized to an adenosine aptamer (in green). Adenosine can induce structure

switching in the aptamer, releasing the short DNA. The device is cycled by adenosine deaminase to turn adenosine into inosine. (d) Inhibition of

thrombin (yellow) activity by a covalently linked thrombin aptamer (green) and activity rescue directed by a specific DNA (in orange) [53�].

Immobilization on a glass surface allows convenient optimal detection. (e) Control of RNA folding by an aptamer [55�]. Two pieces of DNA (in blue)

are covalently attached to an RNA molecule (orange). The addition of a hemin aptamer complementary to the two DNA molecules leads to the

misfolding of the RNA. Hemin can bind to the aptamer and relieve constraints on the RNA to allow it to fold correctly.

multiple cycles. Although aptamers were involved, these

two devices still relied on the energy from DNA hybri-

dization. By introducing a protein enzyme that can che-

mically modify the target molecule of an aptamer, Nutiu

and Li [52��] demonstrated a device free of external

DNA. Initially, a short DNA strand (Figure 4c, in blue)

was hybridized to an adenosine aptamer DNA (in green).

In the presence of adenosine, the DNA in blue was

released and the aptamer DNA bound two adenosine

molecules. The device was cycled by the addition of

adenosine deaminase, which transformed adenosine into

inosine. As the aptamer cannot bind inosine, it re-associ-

ates with the short DNA in blue. Therefore, adenosine

was the fuel for the system and inosine was the waste.

Thrombin activity can be inhibited upon binding to its

aptamer; therefore, controlling aptamer binding to throm-

bin can be used to regulate thrombin activity. Willner and

co-workers [53�] inhibited thrombin by covalently attach-

ing thrombin aptamers to the molecule via a single-

stranded DNA linker (Figure 4d). The addition of

DNA complementary to the linker forced the aptamer

to leave thrombin, thus rescuing enzyme activity. In

another recent study, Miduturu and Silverman [54,55�]


used chemically controlled transitions between DNA

duplexes and aptamer–target complexes to control the

folding of macromolecules. Two short DNA molecules

were covalently attached to the 160-nucleotide

Tetrahymena group I intron P4–P6 RNA domain, which

has well-characterized folding behaviour (Figure 4e). In

the presence of a DNA complementary to the two short

DNAs, RNA folding was constrained because the free

energy favored formation of DNA duplexes. The comple-

mentary DNA also contained a hemin aptamer overhang.

In the presence of hemin, the constraint was released and

the RNA folded into its native state owing to the release of

one DNA duplex. Although performed on an RNA mole-

cule, the same method could be applicable to controlling

the folding of other biopolymers and polymers.

Aptazymes in nanotechnologyDNAzymes can perform chemical modifications on

nucleic acids, while aptamers can bind a broad range of

molecules. A combination of the two has generated a new

class of functional nucleic acids known as allosteric

DNAzymes or aptazymes [56]. Liu and Lu [57] have

employed an adenosine-dependent aptazyme [58], built

on the basis of the Pb2+-specific DNAzyme, to assemble



gold nanoparticles. In the presence of adenosine, the

substrate was cleaved and the assembly inhibited.

ConclusionsIn summary, although their appearance in the literature

has been very recent, functional DNA molecules such as

aptamers, DNAzymes and aptazymes have already found

application in almost every aspect of DNA nanotechnol-

ogy. Therefore, the field of DNA nanotechnology has

been significantly broadened, and the resulting new

materials and devices can penetrate into many other fields

for practical applications, such as sensing, environmental

monitoring, medical diagnostics, drug screening, thera-

peutics, nanoelectronics, nanophotonics, and quantum

computing [18,59,60]. In the future, we are likely to

see the incorporation of individual devices into more

complex multifaceted systems. Multidisciplinary

research, such as the application of functional DNA-

based nanodevices in microfluidics, biophysics and biol-

ogy, will broaden the practical applications of such

devices in many new areas. Importantly, with the power

to combinatorially select functional nucleic acids, these

molecules will become a rich source for developing

nanotechnology.

UpdateYan and co-workers recently prepared two-dimensional

DNA arrays containing fluorophore-labeled thrombin

aptamers, which showed increased fluorescence in the

presence of thrombin [61]. Such a design allows the

generation of programmable sensor arrays that can be

imaged on the surface.

AcknowledgementsResearch by the Lu group reviewed in this article is based upon worksupported by the Department of Energy through an EnvironmentalRemediation Sciences Program (DE-FG02-01ER63179), the NationalScience Foundation through a Nanoscale Science and Engineering Centergrant (DMR-0117792), the US Army Research Laboratory and the USArmy Research Office under grant number DAAD19-03-1-0227, and bythe Illinois Waste Management Research Center under Contract numberHWR04187.

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33. Chang I-H, Tulock JJ, Liu J, Kim W-S, Cannon DM Jr., Lu Y,Bohn PW, Sweedler JV, Cropek DM: Miniaturized lead sensorbased on lead-specific DNAzyme in a nanocapillaryinterconnected microfluidic device. Environ Sci Technol 2005,39:3756-3761.

34. Garibotti AV, Knudsen SM, Ellington AD, Seeman NC: FunctionalDNAzymes organized into two-dimensional arrays. Nano Lett2006, 6:1505-1507.

35. Carmi N, Balkhi HR, Breaker RR: Cleaving DNA with DNA.Proc Natl Acad Sci USA 1998, 95:2233-2237.

36. Chen Y, Wang M, Mao C: An autonomous DNA nanomotorpowered by a DNA enzyme. Angew Chem Int Ed Engl 2004,43:3554-3557.

37.�

Chen Y, Mao C: Putting a brake on an autonomous DNAnanomotor. J Am Chem Soc 2004, 126:8626-8627.

A DNAzyme-based free-running nanomotor was previously reported. Thispaper demonstrated a way to control the free-running nanomotor byusing inactive, but structurally related, DNA sequences.

38.��

Tian Y, He Y, Chen Y, Yin P, Mao C: A DNAzyme that walksprocessively and autonomously along a one-dimensionaltrack. Angew Chem Int Ed Engl 2005, 44:4355-4358.

A sophisticated system was designed to transport a DNAzyme along aDNA track. In this ‘walking’ system, the walker carries an enzymaticfunction that enables it to move forward.

39. Nutiu R, Li Y: Structure-switching signaling aptamers.J Am Chem Soc 2003, 125:4771-4778.

40.��

Nutiu R, Li Y: Structure-switching signaling aptamers:transducing molecular recognition into fluorescencesignalling. Chem Eur J 2004, 10:1868-1876.

Contains a detailed description of how to tune aptamer binding statesbetween the cognate analyte and the complementary DNA.

41. Pavlov V, Xiao Y, Shlyahovsky B, Willner I: Aptamer-functionalized Au nanoparticles for the amplified opticaldetection of thrombin. J Am Chem Soc 2004, 126:11768-11769.

42. Padmanabhan K, Padmanabhan KP, Ferrara JD, Sadler JE,Tulinsky A: The structure of a-thrombin inhibited by a15-mer single-stranded DNA aptamer. J Biol Chem 1993,268:17651-17654.

43.�

Huang C-C, Huang Y-F, Cao Z, Tan W, Chang H-T: Aptamer-modified gold nanoparticles for colorimetric determination ofplatelet-derived growth factors and their receptors. Anal Chem2005, 77:5735-5741.


Platelet-derived growth factors (PDGF) were used to assemble PDGFaptamer-functionalized gold nanoparticles, taking advantage of the factthat each PDGF can bind to two PDGF aptamers. The assembly processwas monitored through the use of a color-change assay.

44.��

Liu J, Lu Y: Fast colorimetric sensing of adenosine andcocaine based on a general sensor design involvingaptamers and nanoparticles. Angew Chem Int Ed Engl 2006,45:90-94.

Two chemical responsive systems were demonstrated that use aptamerDNA to template the assembly of nanoparticles. In the presence of targetmolecules, aggregated nanoparticles disassembled into a dispersedstate in seconds, accompanied by a purple-to-red color change. Suchmaterials are useful for colorimetric sensing of a broad range of analytes.

45.��

Liu J, Lu Y: Smart nanomaterials responsive to multiplechemical stimuli with controllable cooperativity. Adv Mater2006, 18:1667-1671.

Materials whose assembly could be controlled by multiple chemicalswere prepared on the basis of aptamer-linked nanoparticle aggregates;the cooperativity between chemical stimuli can be controlled by differentmaterial designs. Any two combinations of adenosine, cocaine or potas-sium ions were demonstrated to control nanoparticle assembly statesand optical properties.

46.�

Liu Y, Lin C, Li H, Yan H: Aptamer-directed self-assembly ofprotein arrays on a DNA nanostructure. Angew Chem Int EdEngl 2005, 44:4333-4338.

In this study, a thrombin aptamer was incorporated into a DNA periodicstructure. Binding of thrombin to the DNA structure enabled the genera-tion of a thrombin array.

47.�

Levy M, Cater SF, Ellington AD: Quantum-dot aptamer beaconsfor the detection of proteins. ChemBioChem 2005, 6:2163-2166.

A quencher-labeled DNA was positioned close to a quantum dot byhybridization to an attached thrombin aptamer. The addition of thrombinreleased the quencher labeled DNA, unmasking the fluorescence. Thisdevice has been used for sensing applications.

48. Polsky R, Gill R, Kaganovsky L, Willner I: Nucleic acid-functionalized Pt nanoparticles: catalytic labels for theamplified electrochemical detection of biomolecules.Anal Chem 2006, 78:2268-2271.

49. Li JJ, Tan W: A single DNA molecule nanomotor. Nano Lett2002, 2:315-318.

50. Bock LC, Griffin LC, Latham JA, Vermaas EH, Toole JJ: Selectionof single-stranded DNA molecules that bind and inhibit humanthrombin. Nature 1992, 355:564-566.

51.�

Dittmer WU, Reuter A, Simmel FC: A DNA-based machine thatcan cyclically bind and release thrombin. Angew Chem Int EdEngl 2004, 43:3550-3553.

This paper demonstrated the concept of using aptamer-binding to controlthe conformation of DNA. Importantly, the aptamer can be used as a‘nanohand’ to grab and release proteins.

52.��

Nutiu R, Li Y: A DNA-protein nanoengine for ‘on-demand’release and precise delivery of molecules. Angew Chem Int EdEngl 2005, 44:5464-5467.

Most previously reported DNA machines require the use of DNA hybri-dization as an energy source. This paper, however, reports the first DNAmachine to be run with non-DNA fuel. Here, a protein enzyme wasemployed to chemically modify the target of an aptamer thus controllingaptamer conformation in solution, with the target molecule functioning asfuel for the system.

53.�

Pavlov V, Shlyahovsky B, Willner I: Fluorescence detection ofDNA by the catalytic activation of an aptamer/thrombincomplex. J Am Chem Soc 2005, 127:6522-6523.

Thrombin activity was inhibited by a covalently attached DNA aptamer;the activity can be rescued by removal of aptamer binding with com-plementary DNA.

54. Miduturu CV, Silverman SK: DNA constraints allow rationalcontrol of macromolecular conformation. J Am Chem Soc2005, 127:10144-10145.

55.�

Miduturu CV, Silverman SK: Modulation of DNA constraints thatcontrol macromolecular folding. Angew Chem Int Ed Engl 2006,45:1918-1921.

Several methods to control the folding of a DNA-modified RNA moleculewere demonstrated, including the use of a hemin aptamer to control RNAfolding by hemin.



56. Soukup GA, Breaker RR: Nucleic acid molecular switches.Trends Biotechnol 1999, 17:469-476.

57. Liu J, Lu Y: Adenosine-dependent assembly of aptazyme-functionalized gold nanoparticles and its application as acolorimetric biosensor. Anal Chem 2004, 76:1627-1632.

58. Wang DY, Lai BHY, Sen D: A general strategy for effector-mediated control of RNA-cleaving ribozymes and DNAenzymes. J Mol Biol 2002, 318:33-43.


59. Famulok M: Allosteric aptamers and aptazymes as probesfor screening approaches. Curr Opin Mol Ther 2005,7:137-143.

60. Lee JF, Stovall GM, Ellington AD: Aptamer therapeuticsadvance. Curr Opin Chem Biol 2006, 10:282-289.

61. Lin C, Katilius E, Liu Y, Zhang J, Yan H: Self-assembled signalingaptamer DNA arrays for protein detection. Angew Chem Int Ed2006, 45:5296.


functional dna nanotechnology: emerging applications of dnazymes and aptamers

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