Synthetic Polymer Hybridization with DNA and RNA Directs Nanoparticle Loading, Silencing Delivery, and Aptamer Function

We report herein discrete triplex hybridization of DNA and RNA with polyacrylates. Length-monodisperse triazine-derivatized polymers were prepared on gram-scale by reversible addition–fragmentation chain-transfer polymerization. Despite stereoregio backbone heterogeneity, the triazine polymers bind T/U-rich DNA or RNA with nanomolar affinity upon mixing in a 1:1 ratio, as judged by thermal melts, circular dichroism, gel-shift assays, and fluorescence quenching. We call these polyacrylates “bifacial polymer nucleic acids” (bP_oNAs). Nucleic acid hybridization with bP_oNA enables DNA loading onto polymer nanoparticles, siRNA silencing delivery, and can further serve as an allosteric trigger of RNA aptamer function. Thus, bP_oNAs can serve as tools for both non-covalent bioconjugation and structure–function nucleation. It is anticipated that bP_oNAs will have utility in both bio- and nanotechnology

Bifacial Peptide Nucleic Acid Directs Cooperative Folding and Assembly of Binary, Ternary, and Quaternary DNA Complexes

We report herein the structuring of single-stranded thymine-rich DNA sequences into peptide–DNA hairpin triplex structures via designed melamine–thymine nucleobase recognition. Melamine-displaying α-peptides were synthesized with the general form (EM*)_<i>n</i>, where M* denotes a lysine residue side chain derivatized with melamine, a bifacial hydrogen bond complement for thymine. We have found that (EM*)_<i>n</i> peptides, which we term bifacial peptide nucleic acid (bPNA), function as a noncovalent template for thymine-rich DNA tracts. Unstructured DNA of the general form dT_<i>n</i>C_<i>m</i>T_<i>n</i> are bound to (EM*)_<i>n</i> peptides and fold into cooperatively melting 1:1 bPNA–DNA hairpin complexes with dissociation constants in the submicromolar to low nanomolar range for <i>n</i> = 4–10. As the length of the interface (<i>n</i>) is decreased, the melting temperature of the bPNA–DNA complex drops significantly, though <i>K</i>_d increases are less substantial, suggestive of strong enthalpy–entropy compensation. This is borne out by differential scanning calorimetry analysis, which indicates enthalpically driven bPNA–DNA base-stacking that becomes markedly less exothermic as the recognition surface <i>n</i> decreases in size. The recognition interface tolerates a high number of “mismatches” and indicates half-site, or monofacial, recognition between melamine and thymine may occur if only 1 complementary nucleobase is available. Association correlates directly with fractional thymine content, with optimal binding when the number of T–T sites match the number of melamine units. Interestingly, when a DNA host has more T–T sites than melamine sites on bPNA, two or three bPNAs can bind to a single DNA, resulting in ternary and quaternary complexes that have higher thermal stability than the binary (1:1) bPNA–DNA complex, suggestive of cooperative multisite binding. In contrast, when two bPNAs of different lengths bind to the same DNA host, a ternary complex is formed with two melting transitions, corresponding to independent melting of each bPNA component from the complex. These data demonstrate that melamine-displaying bPNA recognize thymine-rich DNA in predictable and multifaceted ways that allow binding affinity, structure stability, and stoichiometry to be tuned through simple bPNA length modification and matching with DNA length. Synthetic bPNA structuring elements may be useful tools for biotechnology

Bifacial Peptide Nucleic Acid as an Allosteric Switch for Aptamer and Ribozyme Function

We demonstrate herein that bifacial peptide nucleic acid (bPNA) hybrid triplexes functionally substitute for duplex DNA or RNA. Structure–function loss in three non-coding nucleic acids was inflicted by replacement of a duplex stem with unstructured oligo-T/U strands, which are bPNA binding sites. Functional rescue was observed on refolding of the oligo-T/U strands into bPNA triplex hybrid stems. Bifacial PNA binding was thus used to allosterically switch-on protein and small-molecule binding in DNA and RNA aptamers, as well as catalytic bond scission in a ribozyme. Duplex stems that support the catalytic site of a minimal type I hammerhead ribozyme were replaced with oligo-U loops, severely crippling or ablating the native RNA splicing function. Refolding of the U-loops into bPNA triplex stems completely restored splicing function in the hybrid system. These studies indicate that bPNA may have general utility as an allosteric trigger for a wide range of functions in non-coding nucleic acids

The salient region.

<p>The salient region.</p

Guanosine-Based Supramolecular Hydrogels with Dynamic Time-Dependent Fluorescence for Information Encryption

Dynamic time-dependent fluorescent encrypted hydrogels provide a highly secure and sophisticated information security strategy. However, challenges still remain in developing new green encrypted hydrogels with simple preparation. Herein, a new series of guanosine (G)-based supramolecular hydrogels with dynamic time-dependent fluorescence rather than traditional fluorescent materials are exquisitely constructed via a one-pot reaction for the first time. Typically, their fluorescence intensity and wavelength can gradually change with the time dimension of more than 1 year, showing promising potential in information encryption–decryption–destruction. A detailed timescale structure characterization combined with the transition-state analysis of the self-assembly process revealed that the G-quartet-based self-assembly in hydrogels has significant endogenous dynamics, which further affects gelation and time-dependent fluorescence through the self-assembly rate. Further combined with the excellent shear thinning, self-healing, and metal ion response properties, the writing, storage, reading, and ″burning after reading″ of information can be successfully realized in a long-term dimension. Therefore, this study illustrates that the ingenious use of functional supramolecular self-assembly building blocks will be a useful strategy for the development of dynamic time-dependent fluorescent encrypted hydrogels

Peptide Ligation and RNA Cleavage via an Abiotic Template Interface

We report herein DNA- and RNA-templated chemical transformation of bifacial peptide nucleic acid (bPNA) fragments directed by an abiotic triplex hybrid interface. Assembly of one bPNA strand with two unstructured oligo T/U strands enables facile insertion of DNA and RNA template sites within partially folded nucleic acids; this template topology is not easily accessed through native base-pairing. Triplex hybridization of reactive bPNA fragments on DNA and RNA templates is shown to catalyze amide bond ligation and controlled bPNA chain extension. RNA-templated oxidative coupling of bPNA fragments is found to result in the emergence of ribozyme cleavage function, thus establishing a connection between engineered and native reaction sites. These data demonstrate the use of new topologies in nucleic acid-templated chemistry that could serve as chemically sensitive DNA and RNA switches

Guanosine-Based Supramolecular Hydrogels with Dynamic Time-Dependent Fluorescence for Information Encryption

Dynamic time-dependent fluorescent encrypted hydrogels provide a highly secure and sophisticated information security strategy. However, challenges still remain in developing new green encrypted hydrogels with simple preparation. Herein, a new series of guanosine (G)-based supramolecular hydrogels with dynamic time-dependent fluorescence rather than traditional fluorescent materials are exquisitely constructed via a one-pot reaction for the first time. Typically, their fluorescence intensity and wavelength can gradually change with the time dimension of more than 1 year, showing promising potential in information encryption–decryption–destruction. A detailed timescale structure characterization combined with the transition-state analysis of the self-assembly process revealed that the G-quartet-based self-assembly in hydrogels has significant endogenous dynamics, which further affects gelation and time-dependent fluorescence through the self-assembly rate. Further combined with the excellent shear thinning, self-healing, and metal ion response properties, the writing, storage, reading, and ″burning after reading″ of information can be successfully realized in a long-term dimension. Therefore, this study illustrates that the ingenious use of functional supramolecular self-assembly building blocks will be a useful strategy for the development of dynamic time-dependent fluorescent encrypted hydrogels

The salient region.

<p>The salient region.</p

Average F-measures obtained from different settings of <i>m</i>.

<p>Average F-measures obtained from different settings of <i>m</i>.</p

Results of segmentation on images with complex backgrounds.

<p>Results of segmentation on images with complex backgrounds.</p