Communication pubs.acs.org/cm
Quantum Dot Encapsulation Using a Peptide-Modified Tetrahedral DNA Cage Divita Mathur,†,⊥,# Anirban Samanta,†,⊥,# Eunkeu Oh,‡,∥ Sebastián A. Díaz,†,∇ Kimihiro Susumu,‡,∥ Mario G. Ancona,§ and Igor L. Medintz*,† †
Center for Bio/Molecular Science and Engineering Code 6900, ‡Optical Sciences Division Code 5600, and §Electronics Science and Technology Division Code 6800, U.S. Naval Research Laboratory, Washington, DC 20375, United States ∥ Sotera Defense Solutions, Inc., Columbia, Maryland 21046, United States ⊥ College of Science, George Mason University, Fairfax, Virginia 22030, United States ∇ American Society for Engineering Education, Washington, DC 20036, United States S Supporting Information *
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because they serve a range of more size/scale-sensitive optical applications. Because many QD applications involve FRET, which occurs only over very short distances (1−10 nm),3 cages to be used with QDs need to have thin walls and to fit snugly to minimize movement. Most caging strategies rely on prefunctionalizing the NP with a single-stranded (ss) DNA tether to allow them to bind to a complement on the cage during formation. DNA functionalization of the QD/NP is typically accomplished by thiol−noble metal or biotin−streptavidin interactions and brings with it the need for both stoichiometric chemistry, where single DNA per NP functionalization is especially tricky to achieve, and subsequent purification/ enrichment.9,13 A tether free approach to capturing QDs in DNA icosahedra constructed from distinct five-way junctions displaying overhangs was recently described.16 This strategy required preligating portions of the DNA to form initial subcomponents of the icosahedra, adding these to excess QDs, and then assembling the final structure with a further hybridization reaction where the QDs were encapsulated stochastically followed by further purification prior to use. Our overarching principal to QD encapsulation is simplicity, with attention paid to all aspects of the process including the cage design, QD attachment, and the encapsulation itself. The scheme we use is summarized in Figure 1. The DNA nanocage is as simple as possible being a tetrahedron that is assembled in a single step using a one-pot annealing reaction13 following Turberfield et al.17 It comprises five oligonucleotides (Table 1), where the T1-pep strand is 5′-end modified with a (His)5peptide tag and the T3-Cy3 is 5′-labeled with a Cy3 dye. Positions for the peptide and Cy3 were derived using the approach of Erben et al.18 such that the spatial orientation of the peptidyl handle is meant to be securely toward the inner cavity of the cage whereas the Cy3 molecule is facing outward. The inward-facing (His)5-peptide tag is meant to capture the QD inside the cage, whereas the Cy3 is designed to detect the presence of the QD optically while not introducing any obstruction inside the cage. Cage dimensions are estimated at 7 nm/side with an internal cavity of ∼5 nm diameter using edge-
he unique photophysical properties of luminescent semiconductor quantum dots (QDs) have given them a continuously growing role in many facets of bionanotechnology.1,2 Along with generalized utility as bright, photostable probes, QDs can act as both excellent donors and acceptors within a variety of electron transfer and, more commonly, Förster resonance energy transfer (FRET) configurations.3,4 Applications exploiting QDs in this manner include biosensors, beacons for subcellular imaging, nanoscale rulers, diagnostics, therapeutics, biocomputing, and components of photonic wires and light harvesting circuitry.3−8 Supporting these applications, a library of bioconjugation chemistries have been developed for attaching all manner of biologicals and other molecules to QDs.9 To enable further development of QD-based nanodevices of higher complexity, precise control over the positioning of QDs, organic dyes, and other functional molecules with respect to one another is critically needed. Although quite sophisticated, current attachment chemistries are incapable of achieving this. The advent of structural DNA nanotechnology has made rational design and self-assembly of a variety of 2- and 3dimensional nanostructures possible.10 When used as scaffolds, such DNA assemblies provide fine control over the spatial arrangement of molecules such as dyes that is hard to achieve in other ways.11 Toward the goal of creating discretely ordered yet complex heteromolecular environments around QDs with far greater control over composition and configuration, we describe a novel method of QD encapsulation within a DNAbased cage. In general, DNA nanocages are drawing attention not only for assembly purposes but also for carrying or releasing cargoes, for forming shaped nanoparticles (NPs) and for providing access to designer synthons for building higherorder structures.10−14 Caging QDs and other NPs in a variety of DNA structures ranging from polyhedra to boxes is still a nascent field but not without precedence as elegantly described by Chandrasekaran and Levchenko.13 Gold NPs (AuNPs) have been the primary material used to date along with magnetic NPs.13,14 Precise control over the AuNPs was achieved by inserting them into DNA nanocages, with the power of this technique illustrated by synthesizing AuNP “molecules” with well-controlled “bond” lengths and angles.15 Precisely incorporating QDs into nanoassemblies is even more important © 2017 American Chemical Society
Received: January 10, 2017 Revised: June 27, 2017 Published: July 5, 2017 5762
DOI: 10.1021/acs.chemmater.7b00108 Chem. Mater. 2017, 29, 5762−5766
Communication
Chemistry of Materials
Figure 2. Characterization of cage self-assembly via agarose gel electrophoresis following one-pot assembly of the oligos at 90 °C for 2 min and ambient cooling to room temperature Gel = 1.5% agarose gel in 1× Tris-acetate EDTA buffer (pH 8.3) with 12.5 mM MgCl2 run at 10 V/cm for 1 h. Lanes, left-to-right: Marker (M), T1-pep oligo, hybridized T2-T5 oligos, assembled DNA cage. Calculated cage MW ∼ 78 500 g/mol or ∼124 bp. Lane 2 (T1-pep) presented with enhanced exposure level.
Figure 1. QD encapsulation within a DNA tetrahedral cage. (a) 3D rendering of the DNA cage, containing a (His)5 peptidyl-tag internally and a Cy3 dye on the external edge. Programmability of the DNA(His)5-tag ensures its spatial orientation toward the inner cavity, increasing the probability of QD encapsulation. (b) CdSe/ZnS core/ shell QD structure with surface functionalizing CL4 ligand. QD attachment to the (His)5-tag on the cage is driven by metal affinity coordination (not to scale). (c) Structure of the CL4 ligand.
to-edge and midsphere geometric estimates. Rather than prefunctionalize the QD with a DNA linker or utilizing stochastic encapsulation, we rely on the (His)5-peptide tag to function as a single-point QD capture element within the cage. Extensive work by us and many others has shown that polyhistidine sequences bind to the surface of CdSe/ZnS core/ shell QDs via metal affinity coordination, Figure 1b.9 This highaffinity (Kd ≈ 1 nM) binding interaction occurs almost spontaneously when both materials are added together assuming no steric constraints.9 We first characterized DNA tetrahedral cage self-assembly using gel electrophoresis (Figure 2). Results show the cage forms with high efficiency and contains insignificant populations of free oligos and higher molecular weight complexes (30% loading directly while also being a slightly more facile, efficient protocol. This is specifically for assembly formats using excess cage relative to QD; other formats and stoichiometries were not explored here but may serve to increase (or decrease) the loading efficiency. Nevertheless, such loading efficiencies begin to approach values where samples may be usable directly without further purification.20 The caged QDs are amenable to purification with a centrifugal filter (Figure S7) with the loss of some product. The ability to isolate a QD inside the cage and control the location of an initial dye relative to it now opens up the opportunity to achieve far more complex spatial control15 and site-specific display on the DNA scaffold. We anticipate that a similar capture strategy may be applicable to many other NP types because a variety of small peptidyl motifs are available that are capable of binding to different metals or other nanoscale materials.12,21 Similarly precise positioning may also be applicable to other types of DNA-based FRET structures22 that seek to incorporate QDs6−9 along with those that seek to add other materials including biologicals such as enzymes or drugs.23
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The authors declare no competing financial interest.
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ACKNOWLEDGMENTS The authors acknowledge ONR, NRL, and the NRL-NSI for financial support.
ASSOCIATED CONTENT
* Supporting Information S
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.chemmater.7b00108. Materials and methods, supporting data (PDF)
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REFERENCES
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AUTHOR INFORMATION
Corresponding Author
*I. L. Medintz. E-mail:
[email protected]. ORCID
Sebastián A. Díaz: 0000-0002-5568-0512 Igor L. Medintz: 0000-0002-8902-4687 Author Contributions #
These authors contributed equally. 5765
DOI: 10.1021/acs.chemmater.7b00108 Chem. Mater. 2017, 29, 5762−5766
Communication
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DOI: 10.1021/acs.chemmater.7b00108 Chem. Mater. 2017, 29, 5762−5766
