DNA as a Molecular Ruler: Interrogation of a Tandem SH2 Domain with Self‐Assembled, Bivalent DNA–Peptide Complexes

Conspectus Living systems have evolved a variety of nanostructures to control the molecular interactions that mediate many functions including the recognition of targets by receptors, the binding of enzymes to substrates, and the regulation of enzymatic activity. Mimicking these structures outside of the cell requires methods that offer nanoscale control over the organization of individual network components. Advances in DNA nanotechnology have enabled the design and fabrication of sophisticated one-, two- and three-dimensional (1D, 2D and 3D) nanostructures that utilize spontaneous and sequence specific DNA hybridization. Compared to other self-assembling biopolymers, DNA nanostructures offer predictable and programmable interactions, and surface features to which other nanoparticles and bio-molecules can be precisely positioned. The ability to control the spatial arrangement of the components while constructing highly-organized networks will lead to various applications of these systems. For example, DNA nanoarrays with surface displays of molecular probes can sense noncovalent hybridization interactions with DNA, RNA, and proteins and covalent chemical reactions. DNA nanostructures can also align external molecules into well-defined arrays, which may improve the resolution of many structural determination methods, such as X-ray diffraction, cryo-EM, NMR, and super-resolution fluorescence. Moreover, by constraining target entities to specific conformations, self-assembled DNA nanostructures can serve as molecular rulers to evaluate conformation-dependent activities. This Account describes the most recent advances in the DNA nanostructure directed assembly of biomolecular networks and explores the possibility of applying this technology to other fields of study. Recently, several reports have demonstrated the DNA nanostructure directed assembly of spatially-interactive biomolecular networks. For example, researchers have constructed synthetic multi-enzyme cascades by organizing the position of the components using DNA nanoscaffolds in vitro, or by utilizing RNA matrices in vivo. These structures display enhanced efficiency compared to the corresponding unstructured enzyme mixtures. Such systems are designed to mimic cellular function, where substrate diffusion between enzymes is facilitated and reactions are catalyzed with high efficiency and specificity. In addition, researchers have assembled multiple choromophores into arrays using a DNA nanoscaffold that optimizes the relative distance between the dyes and their spatial organization. The resulting artificial light harvesting system exhibits efficient cascading energy transfers. Finally, DNA nanostructures have been used as assembly templates to construct nanodevices that execute rationally-designed behaviors, including cargo loading, transportation and route control.

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“…The nanoscaffold displayed the two domain-binding ligands at various distances and flexibilities. 44 …”

Section: Label-free Detection Of Bimolecular Interactionsmentioning

confidence: 99%

Spatially-Interactive Biomolecular Networks Organized by Nucleic Acid Nanostructures

2012

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“…17,18 They also demonstrated that bivalent peptide-displayed duplexes with regular intervals as molecular rulers effectively bound to tandem SH2 domain of Syk kinase. 19 Moreover, Montesarchio and co-workers reported antiviral activity against HIV-1 of glycoconjugated DNA quadruplex. 20 Hamilton and co-workers constructed a pentaplex assembly of phosphocholine (PC)DNA conjugate in the presence of Cs + ions and revealed that the pentaplex assembly of PC binds to human C-reactive protein with 350-fold higher affinity than its monomeric counterpart.…”

Section: ¹1mentioning

confidence: 99%

Construction of Functional Biomaterials by Biomolecular Self-Assembly

Matsuura

2017

Bulletin of the Chemical Society of Japan

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This account discusses construction strategies for various functional biomaterials based on the designed self-assembly of biomolecules. Novel glycoclusters with regular intervals were developed by self-assembly of carbohydrate-conjugated oligodeoxyribonucleotides (ODNs) with the half-sliding complementary ODNs. Complexes of carbohydrate-modified DNA and lectin afforded a new regulation system for gene expression. DNA three-way junctions bearing self-complementary stickyends were self-assembled into nanometer-to-micrometer-sized spherical structures depending on the concentration. The threeway component design was extended to the design of an artificial trigonal peptide conjugate. The trigonal peptide conjugates bearing ¢-sheet-forming peptides or glutathione selfassembled into nano-sized spherical assemblies. Self-assembly of ¢-annulus peptide derived from tomato bushy stunt virus afforded artificial viral capsids, which can encapsulate and be modified with various molecules.

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“…2B). 15,32 Thermal denaturation analyses showed that the peptides had a negligible influence on the stability of the DNA duplexes ( Fig. S1 †).…”

Section: Spatial Screening Of the Recombinant α-Ear Domainmentioning

confidence: 99%

Probing heterobivalent binding to the endocytic AP-2 adaptor complex by DNA-based spatial screening

et al. 2015

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The double helical DNA scaffold offers a unique set of properties, which are particularly useful for studies of multivalency in biomolecular interactions: (i) multivalent ligand displays can be formed upon nucleic acid hybridization in a self-assembly process, which facilitates spatial screening (ii) valency and spatial arrangement of the ligand display can be precisely controlled and (iii) the flexibility of the ligand display can be adjusted by integrating nick sites and unpaired template regions. Herein we describe the use of DNA-based spatial screening for the characterization of the adaptor complex 2 (AP-2), a central interaction hub within the endocytic protein network in clathrin-mediated endocytosis. AP-2 is comprised of a core domain and two, so-called appendage domains, the αand the β2-ear, which associate with cytoplasmatic proteins required for the formation or maturation of clathrin/AP-2 coated pits. Each appendage domain has two binding grooves which recognize distinct peptide motives with micromolar affinity. This provides opportunities for enhanced interactions with protein molecules that contain two (or more) different peptide motives. To determine whether a particular, spatial arrangement of binding motifs is required for high affinity binding we probed the distance-affinity relationships by means of DNA-programmed spatial screening with self-assembled peptide-DNA complexes. By using trimolecular and tetramolecular assemblies two different peptides were positioned in 2-22 nucleotide distance. The binding data obtained with both recombinant protein in well-defined buffer systems and native AP-2 in brain extract suggests that the two binding sites of the AP-2 α-appendage can cooperate to provide up to 40-fold enhancement of affinity compared to the monovalent interaction. The distance between the two recognized peptide motives was less important provided that the DNA duplex segments were connected by flexible, single strand segments. By contrast, the experiments with a more rigid, duplex-spaced assembly revealed marked distance dependencies. Consequences for the function of adaptor proteins are discussed. † Electronic supplementary information (ESI) available: Details on the synthesis of peptides, oligonucleotides, peptide nucleic acids, oligonucleotide-peptide conjugates and recombinant GST-fusion protein, characterization data, and binding assays. See

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DNA as a Molecular Ruler: Interrogation of a Tandem SH2 Domain with Self‐Assembled, Bivalent DNA–Peptide Complexes

Cited by 97 publications

References 56 publications

Spatially-Interactive Biomolecular Networks Organized by Nucleic Acid Nanostructures

Spatially-Interactive Biomolecular Networks Organized by Nucleic Acid Nanostructures

Construction of Functional Biomaterials by Biomolecular Self-Assembly

Probing heterobivalent binding to the endocytic AP-2 adaptor complex by DNA-based spatial screening

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