Article Cite This: Acc. Chem. Res. XXXX, XXX, XXX−XXX
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DNA Templated Metal Nanoclusters: From Emergent Properties to Unique Applications Published as part of the Accounts of Chemical Research special issue “Toward Atomic Precision in Nanoscience”. Yuxiang Chen,† M. Lisa Phipps,† James H. Werner,† Saumen Chakraborty,*,§ and Jennifer S. Martinez*,‡ †
Center for Integrated Nanotechnologies, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, United States Department of Chemistry and Biochemistry, Northern Arizona University, Flagstaff, Arizona 86011, United States § Department of Chemistry and Biochemistry, University of Mississippi, Oxford, Mississippi 38677, United States
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‡
CONSPECTUS: Metal nanoclusters containing a few to several hundred atoms with sizes ranging from sub-nanometer to ∼2 nm occupy an intermediate size regime that bridges larger plasmonic nanoparticles and smaller metal complexes. With strong quantum confinement, metal nanoclusters exhibit molecule-like properties. This Account focuses on noble metal nanoclusters that are synthesized within a single stranded DNA template. Compared to other ligand protected metal nanoclusters, DNA-templated metal nanoclusters manifest intriguing physical and chemical properties that are heavily influenced by the design of DNA templates. For example, DNA-templated silver nanoclusters can show bright fluorescence, tunable emission colors, and enhanced stability by tuning the sequence of the encapsulating DNA template. DNA-templated gold nanoclusters can also serve as excellent cocatalysts, which are integratable with other biocatalysts such as enzymes. In this Account, DNA-templated silver and gold nanoclusters are selected as paradigm systems to showcase their emergent properties and unique applications. We first discuss the DNA-templated silver nanoclusters with a focus on the creation of a complementary palette of emission colors, which has potential applications for multiplex assays. The importance of the DNA template toward enhanced stability of silver nanoclusters is also demonstrated. We then introduce a special class of activable fluorescence probes that are based on the fluorescence turn-on phenomena of DNA-templated silver nanoclusters, which are named nanocluster beacons (NCBs). NCBs have distinct advantages over molecular beacons for nucleic acid detection, and their emission mechanisms are also discussed in detail. We then discuss a universal method of creating novel DNA−silver nanocluster aptamers for protein detection with high specificity. The remainder of the Account is devoted to the DNA-templated gold nanoclusters. We demonstrate that DNA−gold nanoclusters can serve as enhancers for enzymatic reduction of oxygen, which is one of the most important reactions in biofuel cells. Although DNA-templated metal nanoclusters are still in their infancy, we anticipate they will emerge as a new type of functional nanomaterial with wide applications in biology and energy science. Future research will focus on the synthesis of size selected DNA−metal nanoclusters with atomic monodispersity, structural determination of different sized DNA−metal nanoclusters, and establishment of structure−property correlations. Some long-standing mysteries, such as the origin of fluorescence and mechanism for emission color tunability, constitute the central questions regarding the photophysical properties of DNA−metal nanoclusters. On the application side, more studies are required to understand the interaction between nanocluster and biological systems. In the foreseeable future, one can expect that new biosensors, catalysts, and functional devices will be invented based on the intriguing properties of well-designed DNA−metal nanoclusters and their composites. Overall, DNA− metal nanoclusters can add additional spotlights into the highly vibrant field of ligand protected, quantum sized metal nanoclusters. dependent photoluminescence.2−4 As a bridge between molecular materials and nanoparticles, metal nanoclusters have attracted increasing research interest and hold promising applications in optics, catalysis, and biomedicine.5−9 Surface ligands are an indispensable part of metal nanoclusters. Beyond preventing aggregation during solution phase
1. INTRODUCTION Noble metal nanoclusters, containing a few to several hundred metal atoms with sizes ranging from sub-nanometer to ∼2 nm, constitute a special class of nanomaterial.1 Unlike their larger counterparts (e.g., plasmonic nanoparticles), the ultrasmall size induces strong quantum confinement effects, which leads to molecule-like properties such as discrete energy levels with sizable band gap, strong optical absorption characterized by multiple single electron transitions, and size and surface © XXXX American Chemical Society
Received: July 24, 2018
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DOI: 10.1021/acs.accounts.8b00366 Acc. Chem. Res. XXXX, XXX, XXX−XXX
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The first example of metal nanoclusters templated by singlestranded DNA (ssDNA) was discovered by Dickson and Petty in 2004.25 In this seminal work, a cytosine rich, 12-base oligonucleotide was utilized as the template to direct the assembly of silver ions, benefiting from cytosine−Ag + interactions, then reduced by NaBH4 to generate silver nanoclusters in aqueous solution at room temperature. This simple synthetic procedure is one of the highlights of using DNA as a template. Since then, significant research progress has been made in creating DNA templated silver nanoclusters (denoted as DNA−Ag NCs) with superb photophysical properties including high quantum yield, excellent brightness, photostability, and tunable emission colors from visible to near IR.37−43 Combining the aforementioned advantages with small size and potentially low toxicity, DNA metal NCs show great promise as novel diagnostic probes for biomedical applications.5,44−47 In this Account, we focus on DNA templated silver and gold nanoclusters as paradigm systems for functional nanoclusters, owing to their emergent properties and unique applications enabled by the design of DNA template. We first discuss DNA−Ag NCs with a focus on the creation of a complementary palette of emission colors, while demonstrating DNA sequence specificity and the nanocluster nature of these syntheses. Then, we showcase nanocluster functionality with a special class of DNA−Ag NCs, termed nanocluster beacons (NCBs), that can be used as probes for nucleic acid detection and give some insights into their emission mechanism. We then discuss how to create novel DNA−Ag NC aptamers for protein detection. Then, we direct our attention to demonstrating broader metal selectivity with DNA templates in the case of gold, and how those DNA−Au NCs can be assembled into hybrid structures and enhancers for enzymatic reduction of oxygen. Finally, we highlight important areas of discovery so that DNA−metal nanoclusters will continuously offer exciting opportunities for both fundamental studies and practical applications.
synthesis, ligands significantly affect the size,10 electronic and optical properties,11 and assembly behaviors12,13 of the underlying metal nanoclusters and sometimes even impart new functionalities.14 Well-defined metal nanoclusters have been successfully prepared using a wide variety of ligands and templates, including small organic ligands (e.g., thiols, phosphines, and weak acids), dendrimers, and synthetic polymers.15−18 Biomolecules can also act as templates for metal nanoclusters.19,20 With the advent of ab initio design, one can now create protein or DNA objects with tremendous structural diversity, and metal centers and nanomaterials can be further incorporated to impart new functions and assemblies.21−23 While often one associates metal coordination with proteins (e.g., through amino acids), DNA can also coordinate metals. Early Raman studies demonstrated that the nitrogen of nucleobases could coordinate metal ions.24 The proposed primary coordination sites between different metal ions (e.g., gold and silver) and the four DNA nucleobases are illustrated in Figure 1.25−27
2. EMERGENT PROPERTIES OF DNA-TEMPLATED SILVER NANOCLUSTERS 2.1. DNA Templates a Complementary Palette of Nanoclusters with Unique Emissive Colors
Figure 1. Molecular structures of the four DNA nucleobases and their proposed primary metal-coordination sites to gold and silver atoms.
One of the most attractive features of fluorescent DNA−Ag NCs is the easy tunability of their emission colors, based on the proper design of the DNA template. For example, using DNA microarrays, Dickson and co-workers selected five ssDNA sequences to produce NCs with emission colors ranging from visible to near IR, but with varying reaction conditions.28 In our own efforts to create a complementary palette of fluorescent DNA−Ag NCs, we systematically investigated how the oligonucleotide length and base sequence can affect emission colors. Eventually, we selected four ssDNAs with optimized length and sequence, which can template Ag NCs that exhibit narrow emission bands from green to the near IR when excited at common laser excitation wavelengths (Figure 2).30 Significantly, this was the first example showing that a complementary palette of emissive DNA−Ag NCs can be synthesized under identical conditions (i.e., same buffer and same pH), which demonstrated that the DNA sequence was the decisive factor for dictating cluster size and fluorescence properties through sequence specificity.
Together with our colleagues in the nanocluster community, we have studied how DNA can template and assemble nanoclusters and how those nanoclusters can be used for defined applications.25,28−31 As Au and Ag are historically important metals for nanocluster formation and catalysis and are isoelectronic, yet have differences in reactivity and optical phenomena, they were the targets of our focus.32−36 Having synthesized fluorescent nanoclusters with small molecule templates,15 we first asked these fundamental questions about the use of DNA as template: (1) Can DNA template a nanocluster of metal atoms? (2) Can nanoclusters be made of differing size or properties by varying the DNA sequence? (3) Can these nanoclusters support new functionalities (e.g., fluorescence)? (4) Can DNA template nanoclusters with different metals? (5) Finally, can one create hybrid materials with DNA templated nanoclusters, to support catalytic reactions? B
DOI: 10.1021/acs.accounts.8b00366 Acc. Chem. Res. XXXX, XXX, XXX−XXX
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puzzle and to better understand if these were nanoclusters or individual metal atoms strung out on the DNA template, we performed silver K-edge extended X-ray absorption fine structure (EXAFS) studies on the AG1−3 NCs.48 Detailed analysis of the EXAFS data revealed two major findings. First, all three Ag NCs have shorter Ag−Ag bonds as compared to metallic silver, which confirmed, for the first time, that they were nanoclusters of silver as characterized by the existence of direct metal−metal bonds as opposed to DNA−Ag complexes. Second, the DNA templates coordinated with Ag NCs via the Ag−N/O ligations. However, for AG1−3, no obvious correlation between NC size and emission color was observed, which implies a complicated origin of fluorescence in DNA− Ag NCs. It is clear that much work remains to be done to fully understand the intriguing nature of fluorescent DNA−Ag NCs. Recently, Petty et al. reported a comprehensive study on a DNA templated Ag106+ nanocluster, where they suggested that this nanocluster has a metal like core encapsulated by the DNA−Ag+ shell with a weak interaction between the core and shell.42 2.2. DNA Template Evolution Yields a Highly Stable Silver Nanocluster
Figure 2. A fluorescent palette (from green to near IR) of DNA−Ag NCs can be created by tuning the DNA template. Adapted with permission from ref 30. Copyright 2010 The Royal Society of Chemistry.
Silver based nanomaterials are known for their susceptibility to oxidation, which can significantly hamper their application.16 In the realm of DNA−Ag NCs, stability is always a major issue for developing any practical use of these novel materials. Therefore, it is of paramount importance to create fluorescent DNA−Ag NCs with a long shelf life under ambient conditions and those that remain stable when challenged with oxidation and elevated temperatures. Toward this end, we discovered a 31-base DNA sequence (5′-ACCCGAACCTGGGCTACCACCCTTAATCCCC-3′), termed “D”, that can template the formation of a red emitting Ag NC (λex/λem = 535 nm/615 nm) with a fluorescence quantum yield around 30%.31 Unlike other Ag NCs reported previously, the D-Ag NC is exceptionally stable against oxidation and maintained 75% and 31% of its original fluorescence intensity after 3 months and 10 months, respectively, of dark storage at room temperature. In addition, D-Ag NC also shows exceptional thermostability and maintained ∼50% of its initial fluorescence at 80 °C in a thermocycler. To further understand the role of specific DNA bases in controlling the fluorescence and stability of D-Ag NCs, we performed systematic alternation of the D-DNA sequence and length. The deletion of the presumed NC templating sequence (i.e., 5′-CCCTTAATCCCC-3′) at the 3′ end of the D sequence or any randomization of the D sequence eliminates the formation of fluorescent Ag NCs. While changes in the internal position of up to 6 bases from the NC-templating sequence toward the 5′ end have little to no effect in the observed fluorescence, any base deletion at the 5′ proximal side or external side led to significant decrease in NC fluorescence. These observations demonstrated the critical role of length and sequence of DNA bases in dictating the optical properties of Ag NCs.
We termed the green emitting NCs (λex/λem = 460/550 nm) as AG1, orange emitting NCs (λex/λem = 530/600 nm) as AG2, red emitting NCs (λex/λem = 595/650 nm) as AG3, and nearIR emitting NCs (λex/λem = 640/700 nm) as AG4 (Figure 2). The quantum yields for the four NCs were determined to be 0.002 (AG1), 0.1 (AG2), 0.64 (AG3), and 0.52 (AG4), respectively. AG1, AG2, and AG4 NCs have shelf-lives in common with typical DNA−Ag NCs, that is, several weeks at 4 °C. On the other hand, AG3 NC is quite susceptible to oxidation, which limits its shelf life to 2−3 days. It is worth noting that AG4 NC is more stable than the other three NCs under high salt conditions (e.g., 25 mM NaCl), presumably due to the better shielding effects of its longer oligonucleotide template, which may be an important factor to consider for designing DNA−Ag NCs with high stability. We also measured the brightness of AG4 at the single molecule level via fluorescence correlation spectroscopy (FCS).30 Compared with a common organic dye (Cy5)− DNA conjugate, we found that AG4 is brighter than Cy5 at low excitation power (less than ∼103 W cm−2), but dimmer at higher excitation power (greater than ∼103 W cm−2), which is attributed to differences in blinking dynamics under the two excitation conditions. The significantly faster blinking dynamics of AG4, as compared to Cy5, may make it a better fluorescence fluctuation-based probe for studying biophysical phenomena that happen on a tens of microsecond time scale (e.g., protein folding). The photostability of AG4 NC was also compared with Cy5−DNA under the same illumination conditions. It is clear that AG4 NCs demonstrated superior photostability to Cy5 dyes, which underwent noticeable photobleaching in ten consecutive FCS measurements. Although a wide spectrum of emission colors for DNA−Ag NCs have been obtained so far, it remains a major challenge to precisely correlate emission properties with NC size/structure and DNA length/sequence. Toward solving this long-standing C
DOI: 10.1021/acs.accounts.8b00366 Acc. Chem. Res. XXXX, XXX, XXX−XXX
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Figure 3. Schematic illustrations of the fluorescence turn-on phenomena of nanocluster beacons (NCSs) and their detection of DNA. (a, b) Fluorescence turn-on mechanism of NCBs by the enhancer sequence. (c) DNA detection mechanism of NCBs. (d) Micrographs of individual NCBs from samples without and with DNA targets. Adapted with permission from ref 46. Copyright 2010 American Chemical Society.
3. UNIQUE APPLICATIONS OF DNA-TEMPLATED METAL NANOCLUSTERS
sequence or other sequences by design), we were able to use NCBs as a light up probe for the detection of nucleic acids.49 The NCB consists of two components: one component is a dark Ag NC templated by a ssDNA with an overhang hybridization sequence in addition to the NC nucleation sequence; the other component is a ssDNA containing an enhancer sequence and an additional target-based hybridization sequence. Without the presence of target DNA, the NCB remains dark. When the target DNA (e.g., a sequence from the human oncogene Braf) is present, the two components of the NCB bind to the target sequence in juxtaposition. This allows the enhancer sequence to be in close proximity to the Ag NCs and subsequently turn on the fluorescence of the nanocluster (Figure 3c). It is worth noting that the simultaneous binding events of the two NCB components ensures a high specificity toward the target gene. Moreover, complementary single molecule imaging of NCBs on a total internal reflection fluorescence (TIRF) microscope showed that a significant increase in the number of red emitting NCBs can be easily observed (Figure 3d) as compared to the sample without target DNA. These results not only demonstrate that NCBs can be applied to quantitative measurements but also show that NCB can be used for single molecule detection of target nucleic acids. Therefore, the NCBs should have detection limit smaller than 1 pM.46 While originally developed with an enhancer sequence that produces red emission, we subsequently designed a palette of emission colors for NCBs, from the green out to the near IR, using the same low-fluorescence nanocluster and different enhancer sequences.49 To gain more insights into the emission mechanism of NCBs, Goodson and co-workers investigated the emission dynamics of NCBs using an ultrafast, time-resolved fluorescence up-conversion technique.51 They speculated that the intense red emission of NCB, when binding to an enhancer sequence, might come from a surface state, as indicated by its nanosecond lifetime. They also observed a large two-photon absorption cross-section value for the red emissive NCB, which is beneficial for multiphoton imaging. Based on the steady state absorption, emission, and transient absorption and timeresolved emission of NCBs, a four level emission mechanism was proposed (Figure 4). According to this model, the strong red emission comes from the surface states, while the weak green emission originates from states associated with the NC
3.1. DNA−Ag Nanocluster Beacons for Nucleic Acid Detection
Rapid detection and precise quantification of specific nucleic acids play key roles in the arena of diagnostics and therapeutics, such as point-of-care pathogen detection, genotyping, and disease surveillance. Among many probes that were invented for the aforementioned applications, fluorescent “turn-on” probes, which light up in the presence of targets with a high ratio of fluorescence signal to background are particularly appealing since the removal of unbound probes is usually difficult. One of the most widely used fluorescent turn-on probes is called a molecular beacon. A molecular beacon is a hairpin-like ssDNA that lights up when binding to specific nucleic acid targets. However, conventional molecular beacons have intrinsic drawbacks. For instance, the imperfect quenching efficiency of the fluorophore in molecular beacons often leads to high background fluorescence and reduced sensitivity toward target nucleic acids. In addition, the need for dual labeling of fluorophore and quencher to make molecular beacons also increases the cost and lowers the yield and purity. To circumvent the limitations associated with molecular beacons, to provide more sensitive detection of nucleic acid targets down to the single molecule level, and to develop fluorophores that can be easily toggled from one color to another, new types of fluorescent probes are needed that do not rely on Förster resonance energy transfer (FRET) based mechanisms. Taking advantage of the fact that the fluorescent properties of DNA−Ag NCs are highly sensitive to the surrounding environment, our group developed a DNA−Ag NC based turn-on probe that fluoresces upon hybridization to target DNA sequences, named nanocluster beacons (NCBs).46,49,50 The invention of NCBs is based on an intriguing fluorescence turn-on phenomenon. An initially weakly green fluorescent DNA−Ag NC can be transformed into a bright red emitting NC when placed in close proximity to a DNA enhancer sequence (Figure 3a,b). The fluorescence enhancement ratio can exceed 500-fold. Because of such immense fluorescence enhancement effects when putting dark Ag NCs in close proximity to an enhancer sequence (e.g., guanine rich D
DOI: 10.1021/acs.accounts.8b00366 Acc. Chem. Res. XXXX, XXX, XXX−XXX
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3.3. DNA−Ag Nanocluster Aptamers for Protein Detection
DNA-templated silver nanoclusters can be designed for protein detection. Direct conjugation between a fluorescent DNA−Ag NC and antibodies could be one option; however, Ag NCs tend to degrade under conjugation conditions. To overcome this well-known problem, it is highly desirable to create a probe that not only can serve as an intrinsic fluorescence reporter but also can contain the recognition motif that could selectively bind with target biomolecules, eliminating the need for a subsequent bioconjugation step. Toward this goal, we developed a prototypical probe by using an intrinsically fluorescent recognition motif comprised of a DNA aptamer-templated Ag NC, which could provide specific binding and sensitive detection of target protein at the same time.53 Aptamers are nucleic acids (e.g., DNA and RNA) that are selectively evolved to bind a wide variety of targets, such as metal ions, small molecules, proteins, and even cancer cells, with high specificity. Inspired by the unique features of aptamers, we reported the first synthesis of a DNA aptamertemplated Ag NC combining the strong fluorescence of DNA− Ag NC with the high binding specificity and affinity of DNA aptamers toward target proteins. Using DNA aptamertemplated Ag NCs, we demonstrated a new strategy for specific and sensitive detection of proteins in a one pot, purification free manner (Figure 5).
Figure 4. Proposed energy diagram for NCB. Adapted with permission from ref 51. Copyright 2012 The Royal Society of Chemistry.
core, denoted as the B state (Figure 4). When the Ag NC binds to the enhancer sequence, a new electronic state (denoted as C) appears, which is observed in transient absorption as an excited state bleach. The C state directly contributes to the red emission of NCB by highly efficient energy transfer between the C state and the surface states (indicated by the blue arrow in Figure 4). It is also possible that the excited state C itself is the emissive state (or state C is the surface state). 3.2. Chameleon Nanocluster Beacons for Single Nucleotide Polymorphisms Detection
Single nucleotide polymorphisms (SNPs) are common single nucleotide genetic variations; therefore, they serve as biomarkers for disease-related genes. Better diagnosis of SNPs not only enables detection of inheritable diseases but also facilitate the identification of cancer type and development of personalized medicine, which maximizes drug efficiency. However, contemporary methods for detecting SNPs were either time-consuming (e.g., enzymatic reactions) or required sophisticated design algorithms (e.g., hybridization probes), which are not ideal for diagnostics at point-of-care settings. Thus, it was of high interest to design a probe that could discriminate among all four single nucleotide variants (A, C, G, and T) in a rapid and reliable manner. Based on all the aforementioned unique features of NCBs, a new type of NCBs called chameleon nanocluster beacons (cNCBs) that exhibited different emission colors when bound to different SNPs was developed.50 Chameleon nanocluster beacons were developed following the observation that the emission of the NCB reliably shifted from red to orange and red again as the DNA enhancer sequence was positioned closer in alignment to the nanocluster templating sequence (based upon DNA position). As compared to other contemporary SNP detection schemes, SNP detection using cNCB could be designed using the exact same DNA sequence to detect multiple versions of SNPs (e.g., same enhancer, nanocluster templating, and target binding DNA). The target binding sequences are designed such that a SNP on the target caused a frame shift in base pairing of the enhancer or nanocluster templating sequence to the target. That frame shift subtly alters the relative positions of the enhancer and nanocluster, changing the color more red or orange (as much as a 60−70 nm change). Thus, in a cNCB-based assay, the fluorescence emission identifies even a single base change within the SNP and the emission intensity quantitates the amount of target. Later, Yeh, Petty, and co-workers extended this work by creating a palette of cNCB emission colors.52
Figure 5. An intrinsically fluorescent recognition ligand is created as a chimera of DNA−AgNC and DNA-aptamers. Inset fluorescent tubes are the detection reactions when the chimera was subjected to specific (thrombin) and nonspecific proteins (lane 1, aptamer−AgNCs; lane 2, aptamer−AgNCs with thrombin protein; lane 3, aptamer−AgNCs with streptavidin; lane 4, aptamer−AgNCs with PDGF; lane 5, aptamer−AgNCs with BSA.) Adapted with permission from ref 53. Copyright 2011 The Royal Society of Chemistry.
As a proof of concept, an abundant protein in human serum, α-thrombin, was selected as target. Based on a previously reported aptamer that can selectively bind thrombin, a chimera DNA−Ag NC aptamer was created. The chimeric DNA sequence contains two segments: the aptamer sequence for selective binding of thrombin, and the cytosine rich DNA sequence for templating growth of Ag NC (Figure 5). This DNA−Ag NC aptamer serves a dual missionfluorescent label and specific binding motifat the same time. Since the DNA−Ag NC aptamer was synthesized in a single step (i.e., similar to other DNA−Ag NC synthesis) and does not require any covalent attachment of aptamer or protein to a E
DOI: 10.1021/acs.accounts.8b00366 Acc. Chem. Res. XXXX, XXX, XXX−XXX
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Figure 6. Electrochemical properties of DNA−Au NC and BOD−Au NC/SWNT composite. (a) CV and (b) DPV scans of DNA−Au NC. (c) Linear sweep voltammetry (LSV) of BOD−plasmonic Au/SWNT (purple), BOD−DNA/SWNT (blue), BOD/SWNT (black), and BOD−Au NC/SWNT (red) composite materials. Traces and shaded areas represent the average and standard deviations of three replicates. (d) ORR composite preparation steps. Adapted with permission from ref 36. Copyright 2015 American Chemical Society.
fluorophore, it is a simple, cost-efficient method for protein detection. Moreover, this DNA−Ag NC aptamer is bright (∼60% quantum yield) with an emission peak at ∼700 nm. When the target protein, thrombin, is present, the fluorescence of the Ag NC is quenched with a detection limit of 1 nM thrombin (Figure 5, inset). Control experiments with denatured thrombin, other nonspecific proteins, and random DNA sequences did not cause any change in the Ag NC fluorescence (Figure 5, inset), confirming that the quenching is specific and attributable to thrombin recognition by the aptamer DNA sequence.
and accepting electrons. Motivated by the electroactive nature of the Au NCs, we prepared composites of DNA−Au NC, single-walled carbon nanotubes (SWNTs), and BOD as the cathodic enzyme (Figure 6d). The BOD−Au NC/SWNT composite showed an onset potential (Eonset) of 1.165 V vs RHE, compared to that of 1.150 V observed for BOD/SWNT composite alone, indicating that the presence of Au NCs lowers the overpotential of O2 reduction by ∼15 mV (Figure 6c). The observed Eonset was superior to many supported Pt nanomaterials including nanoparticles, nanoclusters, and nanotubes by ∼295−155 mV. In addition, the presence of the cluster also caused ∼1.6-fold increase in electrocatalytic current density. The observed improvement in BOD-catalyzed ORR was specific to the Au NCs and not the plasmonic gold particles nor DNA alone, suggesting the unique role of the DNA−Au NCs in facilitating ET (Figure 6c). Mechanistic studies established a selective 4e− reduction of O2 to H2O, with minimal (∼3%) production of the partially 2e− reduced H2O2 product. The likely mechanism by which the Au NCs enhance the enzyme catalyzed ORR is by acting as mediator of ET between the electrode and the enzyme active site, analogous to conductive wires.
3.4. DNA Templated Gold Nanoclusters for Enhanced Enzymatic Electroreduction of Oxygen
Development of efficient catalysts for clean energy production is important to minimize our dependence on fossil fuels. Toward developing materials for electrocatalytic applications and to investigate whether DNA can also template nanoclusters with metals other than Ag, we recently synthesized a DNA templated gold nanocluster and demonstrated it can facilitate electron transfer (ET) within an enzymatic electroreduction process.36 In enzymatic fuel cells, both oxidation and reduction processes are performed by enzymes. Within the anode, fuel (e.g., glucose) is oxidized by glucose oxidase, while within the cathode multicopper oxidase, for example, bilirubin oxidase (BOD), catalyzes oxygen reduction reaction (ORR). The ORR process is usually limited by slow ET kinetics from the electrode to the enzyme active site, necessitating the need for effective ET mediators. With this in mind, we synthesized a seven-atom DNA−Au NC with a metal nanocluster core diameter of ∼1 nm. X-ray photoelectron spectroscopy (XPS) data show the presence of both Au(I) and Au(0), similar to thiolate-protected Au NCs with mixed valency.1 In addition, XPS suggests that nitrogen atoms of DNA bases are likely ligated to Au NC. Cyclic voltammetry (CV) and differential pulse voltammetry (DPV) measurements of the DNA−Au NC showed two peaks at 155 mV and 210 mV vs Ag/AgCl, corresponding to two sequential one-electron oxidation processes (Figure 6a,b). These results indicated that the Au NC is capable of donating
4. CONCLUSIONS AND FUTURE PERSPECTIVES Due to the great work and talent of the nanocluster community, DNA templated metal nanoclusters are emerging as a new type of functional nanomaterial with promising application in biology, chemistry, materials, and energy science. However, some major areas of discovery exist for these quantum sized materials including the syntheses of size selected DNA-templated nanoclusters with atomic monodispersity. From structure determination and photophysical characterization of the ground and excited states of these nanoclusters, one can begin to better understand their optical properties including whether these nanoclusters follow the jellium model or other determining factors (e.g., nanocluster geometry and ligand effects). One can also extend our work on non-natural DNA bases to create nanoclusters with more tailored properties (e.g., altered fluorescence, enhanced F
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catalysis, and more selective assembly).54 Finally, predictive design and structure−function studies could lead to new types of nanoclusters with novel properties. With deeper understanding about the fundamentals of DNA templated metal nanoclusters, one can expect that new biosensors, catalysts, and functional devices will be developed based on well-designed DNA−metal nanoclusters and their composites.
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REFERENCES
(1) Jin, R.; Zeng, C.; Zhou, M.; Chen, Y. Atomically Precise Colloidal Metal Nanoclusters and Nanoparticles: Fundamentals and Opportunities. Chem. Rev. 2016, 116, 10346−10413. (2) Diez, I.; Ras, R. H. A. Fluorescent Silver Nanoclusters. Nanoscale 2011, 3, 1963−1970. (3) Xu, H.; Suslick, K. S. Water-Soluble Fluorescent Silver Nanoclusters. Adv. Mater. 2010, 22, 1078−1082. (4) Chakraborty, I.; Pradeep, T. Atomically Precise Clusters of Noble Metals: Emerging Link between Atoms and Nanoparticles. Chem. Rev. 2017, 117, 8208−8271. (5) Tao, Y.; Li, M.; Ren, J.; Qu, X. Metal Nanoclusters: Novel Probes for Diagnostic and Therapeutic Applications. Chem. Soc. Rev. 2015, 44, 8636−8663. (6) Choi, S.; Dickson, R. M.; Yu, J. Developing Luminescent Silver Nanodots for Biological Applications. Chem. Soc. Rev. 2012, 41, 1867−1891. (7) Shang, L.; Dong, S.; Nienhaus, G. U. Ultra-Small Fluorescent Metal Nanoclusters: Synthesis and Biological Applications. Nano Today 2011, 6, 401−418. (8) Zhang, L.; Wang, E. Metal Nanoclusters: New Fluorescent Probes for Sensors and Bioimaging. Nano Today 2014, 9, 132−157. (9) Luo, Z.; Zheng, K.; Xie, J. Engineering Ultrasmall Water-Soluble Gold and Silver Nanoclusters for Biomedical Applications. Chem. Commun. 2014, 50, 5143−5155. (10) Chen, Y.; Zeng, C.; Kauffman, D. R.; Jin, R. Tuning the Magic Size of Atomically Precise Gold Nanoclusters via Isomeric Methylbenzenethiols. Nano Lett. 2015, 15, 3603−3609. (11) Tlahuice-Flores, A.; Whetten, R. L.; Jose-Yacaman, M. Ligand Effects on the Structure and the Electronic Optical Properties of Anionic Au25(SR)18 Clusters. J. Phys. Chem. C 2013, 117, 20867− 20875. (12) Zeng, C.; Chen, Y.; Kirschbaum, K.; Lambright, K. J.; Jin, R. Emergence of Hierarchical Structural Complexities in Nanoparticles and Their Assembly. Science 2016, 354, 1580−1584. (13) Yoon, B.; Luedtke, W. D.; Barnett, R. N.; Gao, J.; Desireddy, A.; Conn, B. E.; Bigioni, T.; Landman, U. Hydrogen-Bonded Structure and Mechanical Chiral Response of a Silver Nanoparticle Superlattice. Nat. Mater. 2014, 13, 807−811. (14) Marjomäki, V.; Lahtinen, T.; Martikainen, M.; Koivisto, J.; Malola, S.; Salorinne, K.; Pettersson, M.; Häkkinen, H. Site-Specific Targeting of Enterovirus Capsid by Functionalized Monodisperse Gold Nanoclusters. Proc. Natl. Acad. Sci. U. S. A. 2014, 111, 1277− 1281. (15) Bao, Y.; Zhong, C.; Vu, D. M.; Temirov, J. P.; Dyer, R. B.; Martinez, J. S. Nanoparticle-Free Synthesis of Fluorescent Gold Nanoclusters at Physiological Temperature. J. Phys. Chem. C 2007, 111, 12194−12198. (16) Desireddy, A.; Conn, B. E.; Guo, J.; Yoon, B.; Barnett, R. N.; Monahan, B. M.; Kirschbaum, K.; Griffith, W. P.; Whetten, R. L.; Landman, U.; Bigioni, T. P. Ultrastable Silver Nanoparticles. Nature 2013, 501, 399−402. (17) Bao, Y.; Yeh, H.-C.; Zhong, C.; Ivanov, S. A.; Sharma, J. K.; Neidig, M. L.; Vu, D. M.; Shreve, A. P.; Dyer, R. B.; Werner, J. H.; Martinez, J. S. Formation and Stabilization of Fluorescent Gold Nanoclusters Using Small Molecules. J. Phys. Chem. C 2010, 114, 15879−15882. (18) Zheng, J.; Dickson, R. M. Individual Water-Soluble DendrimerEncapsulated Silver Nanodot Fluorescence. J. Am. Chem. Soc. 2002, 124, 13982−13983. (19) Xie, J.; Zheng, Y.; Ying, J. Y. Protein-Directed Synthesis of Highly Fluorescent Gold Nanoclusters. J. Am. Chem. Soc. 2009, 131, 888−889. (20) Yang, L.; Yao, C.; Li, F.; Dong, Y.; Zhang, Z.; Yang, D. Synthesis of Branched DNA Scaffolded Super-Nanoclusters with Enhanced Antibacterial Performance. Small 2018, 14, 1800185. (21) Salgado, E. N.; Radford, R. J.; Tezcan, F. A. Metal-Directed Protein Self-Assembly. Acc. Chem. Res. 2010, 43, 661−672.
AUTHOR INFORMATION
Corresponding Authors
*E-mail:
[email protected]. *E-mail:
[email protected]. ORCID
Yuxiang Chen: 0000-0003-1972-9208 James H. Werner: 0000-0002-7616-8913 Saumen Chakraborty: 0000-0002-9256-2769 Jennifer S. Martinez: 0000-0001-6737-2712 Notes
The authors declare no competing financial interest. Biographies Yuxiang Chen is a Director’s Postdoctoral Fellow and University of California/LANL Entrepreneurial Fellow at Los Alamos National Laboratory. He received his Ph.D. in chemistry from Carnegie Mellon University under the supervision of Prof. Rongchao Jin and his B.S. in chemistry from Nankai University, China. His research interests include the design and application of functional composite materials. M. Lisa Phipps is a research technologist within the Center for Integrated Nanotechnologies (CINT) at Los Alamos National Laboratory. She received her B.A. from Colby College and an M.F.A. from San Francisco Art Institute. Her research interests include nanomaterials synthesis and assembly, tissue engineering, and the interaction of materials with cellular systems. James H. Werner is currently the Deputy Group Leader at CINT. He received his bachelor’s degree in applied physics from Caltech and his Ph.D. degree in applied physics from Cornell University, where he was a Hertz fellow. His research interests include instrument development, nanoscience and nanotechnology, and biophysical and analytical applications of single molecule spectroscopy. Saumen Chakraborty is an Assistant Professor at the Chemistry and Biochemistry Department of the University of Mississippi. He earned his Ph.D. in chemistry from the University of Michigan and received postdoctoral training at the University of Illinois and the Los Alamos National Laboratory. His research interests are bioinorganic chemistry and alternate energy research. Jennifer S. Martinez is Professor of Chemistry and Biochemistry and the materials institute, MIRA, at Northern Arizona University. She earned her Ph.D. in chemistry from University of California Santa Barbara. Her research interests are nanocluster synthesis and application, genetically encoded materials, and study of materials interfaces.
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ACKNOWLEDGMENTS The authors acknowledge the Laboratory Directed Research and Development (LDRD) program for a fellowship to Y.C. This work was performed at the Center for Integrated Nanotechnologies, an Office of Science User Facility operated for the U.S. Department of Energy (DOE) Office of Science. G
DOI: 10.1021/acs.accounts.8b00366 Acc. Chem. Res. XXXX, XXX, XXX−XXX
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Accounts of Chemical Research
Cluster within an Encapsulating DNA Host. J. Am. Chem. Soc. 2016, 138, 3469−3477. (43) Thyrhaug, E.; Bogh, S. A.; Carro-Temboury, M. R.; Madsen, C. S.; Vosch, T.; Zigmantas, D. Ultrafast Coherence Transfer in DNATemplated Silver Nanoclusters. Nat. Commun. 2017, 8, 15577. (44) Obliosca, J. M.; Liu, C.; Yeh, H.-C. Fluorescent Silver Nanoclusters as DNA Probes. Nanoscale 2013, 5, 8443−8461. (45) Guo, W.; Yuan, J.; Dong, Q.; Wang, E. Highly SequenceDependent Formation of Fluorescent Silver Nanoclusters in Hybridized DNA Duplexes for Single Nucleotide Mutation Identification. J. Am. Chem. Soc. 2010, 132, 932−934. (46) Yeh, H.-C.; Sharma, J.; Han, J. J.; Martinez, J. S.; Werner, J. H. A DNA−Silver Nanocluster Probe That Fluoresces upon Hybridization. Nano Lett. 2010, 10, 3106−3110. (47) Liu, X.; Wang, F.; Aizen, R.; Yehezkeli, O.; Willner, I. Graphene Oxide/Nucleic-Acid-Stabilized Silver Nanoclusters: Functional Hybrid Materials for Optical Aptamer Sensing and Multiplexed Analysis of Pathogenic DNAs. J. Am. Chem. Soc. 2013, 135, 11832−11839. (48) Neidig, M. L.; Sharma, J.; Yeh, H.-C.; Martinez, J. S.; Conradson, S. D.; Shreve, A. P. Ag K-Edge EXAFS Analysis of DNATemplated Fluorescent Silver Nanoclusters: Insight into the Structural Origins of Emission Tuning by DNA Sequence Variations. J. Am. Chem. Soc. 2011, 133, 11837−11839. (49) Yeh, H. C.; Sharma, J.; Han, J. J.; Martinez, J. S.; Werner, J. H. A Beacon of Light. IEEE Nanotechnol. Mag. 2011, 5, 28−33. (50) Yeh, H.-C.; Sharma, J.; Shih, I.-M.; Vu, D. M.; Martinez, J. S.; Werner, J. H. A Fluorescence Light-Up Ag Nanocluster Probe That Discriminates Single-Nucleotide Variants by Emission Color. J. Am. Chem. Soc. 2012, 134, 11550−11558. (51) Yau, S. H.; Abeyasinghe, N.; Orr, M.; Upton, L.; Varnavski, O.; Werner, J. H.; Yeh, H.-C.; Sharma, J.; Shreve, A. P.; Martinez, J. S.; Goodson, T., III Bright Two-Photon Emission and Ultra-Fast Relaxation Dynamics in a DNA-Templated Nanocluster Investigated by Ultra-Fast Spectroscopy. Nanoscale 2012, 4, 4247−4254. (52) Obliosca, J. M.; Babin, M. C.; Liu, C.; Liu, Y.-L.; Chen, Y.-A.; Batson, R. A.; Ganguly, M.; Petty, J. T.; Yeh, H.-C. A Complementary Palette of NanoCluster Beacons. ACS Nano 2014, 8, 10150−10160. (53) Sharma, J.; Yeh, H.-C.; Yoo, H.; Werner, J. H.; Martinez, J. S. Silver Nanocluster Aptamers: In Situ Generation of Intrinsically Fluorescent Recognition Ligands for Protein Detection. Chem. Commun. 2011, 47, 2294−2296. (54) Chakraborty, S.; Rocha, R. C.; Desireddy, A.; Artyushkova, K.; Sanchez, T. C.; Perry, A. T.; Atanassov, P.; Martinez, J. S. Gold Nanocluster Formation using Morpholino Oligomer as Template and Assembly Agent within Hybrid Bio-Nanomaterials″. RSC Adv. 2016, 6 (93), 90624−90630.
(22) Jones, M. R.; Seeman, N. C.; Mirkin, C. A. Programmable materials and the nature of the DNA bond. Science 2015, 347, 1260901. (23) Yang, D.; Hartman, M. R.; Derrien, T. L.; Hamada, S.; An, D.; Yancey, K. G.; Cheng, R.; Ma, M.; Luo, D. DNA Materials: Bridging Nanotechnology and Biotechnology. Acc. Chem. Res. 2014, 47, 1902− 1911. (24) Duguid, J.; Bloomfield, V. A.; Benevides, J.; Thomas, G. J. Raman Spectroscopy of DNA-Metal Complexes. I. Interactions and Conformational Effects of the Divalent Cations: Mg, Ca, Sr, Ba, Mn, Co, Ni, Cu, Pd, and Cd. Biophys. J. 1993, 65, 1916−1928. (25) Petty, J. T.; Zheng, J.; Hud, N. V.; Dickson, R. M. DNATemplated Ag Nanocluster Formation. J. Am. Chem. Soc. 2004, 126, 5207−5212. (26) Liu, J. DNA-Stabilized, Fluorescent, Metal Nanoclusters for Biosensor Development. TrAC, Trends Anal. Chem. 2014, 58, 99− 111. (27) New, S. Y.; Lee, S. T.; Su, X. D. DNA-Templated Silver Nanoclusters: Structural Correlation and Fluorescence Modulation. Nanoscale 2016, 8, 17729−17746. (28) Richards, C. I.; Choi, S.; Hsiang, J.-C.; Antoku, Y.; Vosch, T.; Bongiorno, A.; Tzeng, Y.-L.; Dickson, R. M. OligonucleotideStabilized Ag Nanocluster Fluorophores. J. Am. Chem. Soc. 2008, 130, 5038−5039. (29) Gwinn, E. G.; O’Neill, P.; Guerrero, A. J.; Bouwmeester, D.; Fygenson, D. K. Sequence-Dependent Fluorescence of DNA-Hosted Silver Nanoclusters. Adv. Mater. 2008, 20, 279−283. (30) Sharma, J.; Yeh, H.-C.; Yoo, H.; Werner, J. H.; Martinez, J. S. A Complementary Palette of Fluorescent Silver Nanoclusters. Chem. Commun. 2010, 46, 3280−3282. (31) Sharma, J.; Rocha, R. C.; Phipps, M. L.; Yeh, H.-C.; Balatsky, K. A.; Vu, D. M.; Shreve, A. P.; Werner, J. H.; Martinez, J. S. A DNATemplated Fluorescent Silver Nanocluster with Enhanced Stability. Nanoscale 2012, 4, 4107−4110. (32) Ritchie, C. M.; Johnsen, K. R.; Kiser, J. R.; Antoku, Y.; Dickson, R. M.; Petty, J. T. Ag Nanocluster Formation Using a Cytosine Oligonucleotide Template. J. Phys. Chem. C 2007, 111, 175−181. (33) Liu, G.; Shao, Y.; Ma, K.; Cui, Q.; Wu, F.; Xu, S. Synthesis of DNA-Templated Fluorescent Gold Nanoclusters. Gold Bull. 2012, 45, 69−74. (34) Liu, G.; Yong, S.; Fei, W.; Shujuan, X.; Jian, P.; Lingling, L. DNA-Hosted Fluorescent Gold Nanoclusters: Sequence-Dependent Formation. Nanotechnology 2013, 24, No. 015503. (35) Kennedy, T. A. C.; MacLean, J. L.; Liu, J. Blue Emitting Gold Nanoclusters Templated by Poly-Cytosine DNA at Low pH and PolyAdenine DNA at Neutral pH. Chem. Commun. 2012, 48, 6845−6847. (36) Chakraborty, S.; Babanova, S.; Rocha, R. C.; Desireddy, A.; Artyushkova, K.; Boncella, A. E.; Atanassov, P.; Martinez, J. S. A Hybrid DNA-Templated Gold Nanocluster For Enhanced Enzymatic Reduction of Oxygen. J. Am. Chem. Soc. 2015, 137, 11678−11687. (37) Vosch, T.; Antoku, Y.; Hsiang, J.-C.; Richards, C. I.; Gonzalez, J. I.; Dickson, R. M. Strongly Emissive Individual DNA-Encapsulated Ag Nanoclusters as Single-Molecule Fluorophores. Proc. Natl. Acad. Sci. U. S. A. 2007, 104, 12616−12621. (38) Schultz, D.; Gardner, K.; Oemrawsingh, S. S. R.; Markešević, N.; Olsson, K.; Debord, M.; Bouwmeester, D.; Gwinn, E. Evidence for Rod-Shaped DNA-Stabilized Silver Nanocluster Emitters. Adv. Mater. 2013, 25, 2797−2803. (39) Petty, J. T.; Story, S. P.; Hsiang, J.-C.; Dickson, R. M. DNATemplated Molecular Silver Fluorophores. J. Phys. Chem. Lett. 2013, 4, 1148−1155. (40) Latorre, A.; Somoza, Á . DNA-Mediated Silver Nanoclusters: Synthesis, Properties and Applications. ChemBioChem 2012, 13, 951− 958. (41) Yuan, Z.; Chen, Y.-C.; Li, H.-W.; Chang, H.-T. Fluorescent Silver Nanoclusters Stabilized by DNA Scaffolds. Chem. Commun. 2014, 50, 9800−9815. (42) Petty, J. T.; Sergev, O. O.; Ganguly, M.; Rankine, I. J.; Chevrier, D. M.; Zhang, P. A Segregated, Partially Oxidized, and Compact Ag10 H
DOI: 10.1021/acs.accounts.8b00366 Acc. Chem. Res. XXXX, XXX, XXX−XXX
