With the introduction of the DNA origami technique, it became possible to rapidly synthesize almost arbitrarily shaped molecular nanostructures at nearly stoichiometric yields. The technique furthermore provides absolute addressability in the sub-nm range, rendering DNA origami nanostructures highly attractive substrates for the controlled arrangement of functional species such as proteins, dyes, and nanoparticles. Consequently, DNAorigami nanostructures have found applications in numerous areas of fundamental and applied research, ranging from drug delivery to biosensing to plasmonics to inorganic materials synthesis. Since many of those applications rely on structurally intact, well-definedDNA origami shapes, the issue of DNA origami stability under numerous application-relevant environmental conditions has received increasing interest in the past few years. In this mini-review we discuss the structural stability, denaturation, and degradation of DNA origami nanostructures under different conditions relevant to the fields of biophysics and biochemistry, biomedicine, and materials science, and the methods to improve their stability for desired applications.
DNA origami represent powerful platforms for single-molecule investigations of biomolecular processes. The required structural integrity of the DNA origami may, however, pose significant limitations regarding their applicability, for instance in protein folding studies that require strongly denaturing conditions. Here, we therefore report a detailed study on the stability of 2D DNA origami triangles in the presence of the strong chaotropic denaturing agents urea and guanidinium chloride (GdmCl) and its dependence on concentration and temperature. At room temperature, the DNA origami triangles are stable up to at least 24 h in both denaturants at concentrations as high as 6 M. At elevated temperatures, however, structural stability is governed by variations in the melting temperature of the individual staple strands. Therefore, the global melting temperature of the DNA origami does not represent an accurate measure of their structural stability. Although GdmCl has a stronger effect on the global melting temperature, its attack results in less structural damage than observed for urea under equivalent conditions. This enhanced structural stability most likely originates from the ionic nature of GdmCl. By rational design of the arrangement and lengths of the individual staple strands used for the folding of a particular shape, however, the structural stability of DNA origami may be enhanced even further to meet individual experimental requirements. Overall, their high stability renders DNA origami promising platforms for biomolecular studies in the presence of chaotropic agents, including single-molecule protein folding or structural switching.
GuidoG rundmeier, [a] AdrianK eller,* [a] andV eikkoL inko* [a, b, c] DNA nanostructures have emerged as intriguing tools for numerous biomedical applications. However, in many of those applicationsa nd most notably in drug delivery,t heir stability and function may be compromised by the biological media. A particularly important issue for medicala pplications is their interaction with proteins such as endonucleases, which may degrade the well-defined nanoscale shapes.H erein, fundamental insights into this interaction are provided by monitoring DNase Id igestion of four structurally distinct DNA origami nanostructures (DONs) in real time and at as ingle-structure level by using high-speed atomic force microscopy.T he effect of the solid-liquidi nterface on DON digestioni sa lso assessed by comparison with experiments in bulk solution. It is shown that DON digestion is strongly dependentoni ts superstructure and flexibility and on the local topology of the individual structure.The rapidly evolvingf ield of DNA nanotechnology enables custom fabrication of various nanoscale shapes with unprecedented addressability; [1] these have found some fascinating implementations in materials science and especiallyi nm any biochemicala nd biophysical systems. [2][3][4][5] In recenty ears, programmable DNA origami nanostructures (DONs) [6][7][8][9][10] have been considered promising candidates for the development of tailored and multifunctional drug-delivery vehicles. [11][12][13][14] For therapeutic in vivo applications, the DON vehicles should preferably be compatible with the immunes ystem, resistant to nucleases, have as ufficiently long circulation half-life, and be able to maintain their shape at the ionic strengths of the biological fluids. [15] However,c oncerns have been raised regarding the stabilityo fD ONs and their performance in biological media. [16, 17] Although it has been shown that DONs can survive under low-cation conditions, [18,19] stability studies performedi n serum or cell culture media have yielded somewhat controversial and quite distinct results. [8,15,[19][20][21] Therefore, significant efforts have been madei nto coating ands tabilizing DONs under biologically relevant conditions. [17,[22][23][24][25] Herein, we study DON digestion by endonucleases (DNase I) in dependence of DON superstructure. DNase Ii sw idely present in serum and varioust issues,w hich makes it one of the most relevantt hreatst ot he stability of DONs in vivo. Previously,D ON cleavage by endonucleases was studied by employing ratherl ong timescales. [8,16] Therefore, thesea pproaches can only resolve the time points at which the whole or most of the nanostructure has been digested. Moreover,t hese experiments could neither reveals patial variations of DNase Is usceptibility within as ingle DON, nor facilitate parallel comparison of different structures under the same conditions. Herein, we thus employ high-speed atomicf orce microscopy (HS-AFM) [26] to study the degradation of four well-established and structurally disti...
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