Exceptional Nuclease Resistance of Paranemic Crossover (PX) DNA and Crossover-Dependent Biostability of DNA Motifs.

Nanometer-sized features and molecular recognition properties make DNA a useful material for nanoscale construction, but degradation in biological fluids poses a considerable roadblock to biomedical applications of DNA nanotechnology. Here, we report the remarkable biostability of a multistranded motif called paranemic crossover (PX) DNA. Compared to double stranded DNA, PX DNA has dramatically enhanced (sometimes >1000 fold) resistance to degradation by four different nucleases, bovine and human serum, and human urine. We trace the cause of PX's biostability to DNA crossovers, showing a continuum of protection that scales with the number of crossovers. These results suggest that enhanced biostability can be engineered into DNA nanostructures by adopting PX-based architectures or by strategic crossover placement.

[1]  David A Rusling,et al.  Triplex-forming oligonucleotides: a third strand for DNA nanotechnology , 2017, Nucleic acids research.

[2]  N. Seeman,et al.  A robust DNA mechanical device controlled by hybridization topology , 2002, Nature.

[3]  Shunqing Zhang,et al.  DNA Nanocarriers: Programmed to Deliver. , 2018, Trends in biochemical sciences.

[4]  P. Rothemund.  Folding DNA to create nanoscale shapes and patterns , 2006, Nature.

[5]  Hari K. K. Subramanian,et al.  Covalent Linkage of One-Dimensional DNA Arrays Bonded by Paranemic Cohesion. , 2015, ACS nano.

[6]  Daniela Rhodes,et al.  G-quadruplex structures: in vivo evidence and function. , 2009, Trends in cell biology.

[7]  William M. Shih,et al.  Virus-Inspired Membrane Encapsulation of DNA Nanostructures To Achieve In Vivo Stability , 2014, ACS nano.

[8]  Prabal K Maiti,et al.  The stability of Seeman JX DNA topoisomers of paranemic crossover (PX) molecules as a function of crossover number. , 2004, Nucleic acids research.

[9]  H. Sleiman,et al.  DNA nanostructure serum stability: greater than the sum of its parts. , 2013, Chemical communications.

[10]  William M. Shih,et al.  Addressing the Instability of DNA Nanostructures in Tissue Culture , 2014, ACS nano.

[11]  N. Seeman,et al.  The unusual and dynamic character of PX-DNA , 2015, Nucleic acids research.

[12]  N. Seeman,et al.  Thermodynamics of forming a parallel DNA crossover. , 2009, Biophysical journal.

[13]  David J. Mooney,et al.  Oligolysine-based coating protects DNA nanostructures from low-salt denaturation and nuclease degradation , 2017, Nature Communications.

[14]  N. Seeman,et al.  Double-stranded DNA homology produces a physical signature , 2010, Proceedings of the National Academy of Sciences.

[15]  M. Laskowski.  12 Deoxyribonuclease I , 1971 .

[16]  C. Mao,et al.  DNA nanotechnology. , 2004, BioTechniques.

[17]  Ruojie Sha,et al.  PX DNA triangle oligomerized using a novel three-domain motif. , 2008, Nano letters.

[18]  N. Seeman,et al.  Paranemic crossover DNA: a generalized Holliday structure with applications in nanotechnology. , 2004, Journal of the American Chemical Society.

[19]  Anindya Dutta,et al.  miR-26a is required for skeletal muscle differentiation and regeneration in mice. , 2012, Genes & development.

[20]  D. Bang,et al.  Self-assembled mirror DNA nanostructures for tumor-specific delivery of anticancer drugs. , 2016, Journal of controlled release : official journal of the Controlled Release Society.

[21]  N. Seeman,et al.  DNase I cleavage of branched DNA molecules. , 1989, The Journal of biological chemistry.

[22]  A Klug,et al.  Sequence-dependent variation in the conformation of DNA. , 1981, Journal of molecular biology.

[23]  Anindya Dutta,et al.  miR-206 and -486 Induce Myoblast Differentiation by Downregulating Pax7 , 2010, Molecular and Cellular Biology.

[24]  Hao Yan,et al.  Single-stranded DNA and RNA origami , 2017, Science.

[25]  N. Stephanopoulos.  Strategies for Stabilizing DNA Nanostructures to Biological Conditions , 2019, Chembiochem : a European journal of chemical biology.

[26]  D. Suck,et al.  X-ray structure of the DNase I-d(GGTATACC)2 complex at 2.3 A resolution. , 1992, Journal of molecular biology.

[27]  D. Suck,et al.  DNA recognition by DNase I , 1994, Journal of molecular recognition : JMR.

[28]  Hélder A. Santos,et al.  Protein Coating of DNA Nanostructures for Enhanced Stability and Immunocompatibility , 2017, Advanced healthcare materials.

[29]  C. Mao,et al.  The study of the paranemic crossover (PX) motif in the context of self-assembly of DNA 2D crystals. , 2016, Organic & biomolecular chemistry.

[30]  Banani Chakraborty,et al.  A Programmable Transducer Self-Assembled from DNA. , 2012, Chemical science.

[31]  Jung-Won Keum,et al.  Enhanced resistance of DNA nanostructures to enzymatic digestion. , 2009, Chemical communications.

[32]  T. Ceska,et al.  Structure-specific DNA cleavage by 5' nucleases. , 1998, Trends in biochemical sciences.

[33]  D. Meldrum,et al.  Stability of DNA origami nanoarrays in cell lysate. , 2011, Nano letters.

[34]  P. Maiti,et al.  Structural rigidity of paranemic crossover and juxtapose DNA nanostructures. , 2011, Biophysical journal.

[35]  Tim Liedl,et al.  One-Step Formation of "Chain-Armor"-Stabilized DNA Nanostructures. , 2015, Angewandte Chemie.

[36]  Baoquan Ding,et al.  Enhanced Stability of DNA Nanostructures by Incorporation of Unnatural Base Pairs. , 2017, Chemphyschem : a European journal of chemical physics and physical chemistry.

[37]  N. Seeman,et al.  Paranemic Crossover DNA: There and Back Again. , 2018, Chemical reviews.

[38]  N. Seeman,et al.  A Proximity-Based Programmable DNA Nanoscale Assembly Line , 2010, Nature.

[39]  N. Seeman,et al.  DNA double-crossover molecules. , 1993, Biochemistry.

[40]  Benjamin D. Walcott,et al.  Complex between a Multicrossover DNA Nanostructure, PX-DNA, and T7 Endonuclease I. , 2019, Biochemistry.

[41]  Michael Matthies,et al.  Block Copolymer Micellization as a Protection Strategy for DNA Origami. , 2017, Angewandte Chemie.

[42]  The PX Motif of DNA Binds Specifically to Escherichia coli DNA Polymerase I. , 2019, Biochemistry.

[43]  William M. Shih,et al.  Glutaraldehyde crosslinking of oligolysines coating DNA origami greatly reduces susceptibility to nuclease degradation. , 2020, Journal of the American Chemical Society.

[44]  William M. Shih,et al.  A 1.7-kilobase single-stranded DNA that folds into a nanoscale octahedron , 2004, Nature.