Highly Integrated, Biostable, and Self-Powered DNA Motor Enabling Autonomous Operation in Living Bodies

An ultimate goal of synthetic DNA motor studies is to mimic natural protein motors in biological systems. Here, we rationally designed a highly integrated and biostable DNA motor system with high potential for living body operation, through simple assembly of a Mn2+-dependent DNAzyme-powered DNA motor with a degradable MnO2 nanosheet. The motor system shows outstanding high integration and improved biostability. High integration confers the motor system with the ability to deliver all the core components to the target sites as a whole, thus, enabling precise control of the spatiotemporal distribution of these components and achieving high local concentrations. At the target sites, reduction of the MnO2 nanosheet by intracellular glutathione (GSH) not only releases the DNA motor, which can then be initiated by the intracellular target, but also produces Mn2+ in situ to power the autonomous and progressive operation of the DNA motor. Interestingly, the resultant consumption of GSH in turn protects the DNA motor from destruction by physiological GSH, thus, conferring our motor system with improved biostability, reduced false-positive outputs, and consequently, an increased potential to be applied in a living body. As a proof of concept, the highly integrated DNA motor system was demonstrated to work well for amplified imaging detection of survivin mRNA (mRNA), an important tumor biomarker, in both living cancer cells and living tumor-bearing mice. This work reveals concepts and strategies promoting synthetic DNA motor applications in biological systems

Lignin-Based Hydrogen-Bonded Covalent Organic Polymers as Functional “Switches” of Modified Atmosphere Packaging Membranes for Preservation of Perishable Foods

To reduce poverty and hunger in economically backward countries and regions, increasing demand for addressing severe worldwide food deterioration and wastage has triggered intensive research efforts toward modified atmosphere packaging (MAP) technology. Herein, we demonstrate that natural lignin can be used as a cheap and green monomer for preparation of hydrogen-bonded covalent organic polymers (HCOPs) with good crystallinity, permanent porosity, and high thermostability. The optimal lignin-based HCOPs (named LT-HCOPs) are synthesized by using tetrafluoroterephthalonitrile as the cross-linker, ethanol/water (7/3, v/v) as the reaction solvent, 120 °C as the reaction temperature, and ammonium persulphate and anhydrous potassium carbonate as catalysts. As-prepared LT-HCOPs can be used as promising “switches” for regulating gas permeability in the passive MAP membranes. The LT-HCOP-based passive MAP membrane, which is prepared by a simple mixing and solvent evaporation process, shows good biocompatibility, antimicrobial and antioxidant activities, improved ultraviolet light barrier property, high CO2/O2 selectivity, and suitable H2O permeation and thus is demonstrated to work well in prolonging the shelf life of perishable foods such as strawberries, waxberries, cherries, cherry tomatoes, and mangoes. This work provides a feasible way to use cheap and green natural materials to construct advanced functional materials for practical applications

DNA-Functionalized Porphyrinic Metal–Organic Framework-Based Drug Delivery System for Targeted Bimodal Cancer Therapy

A DNA-functionalized porphyrinic MOF (porMOF) drug delivery system was successfully constructed. porMOF as a photosensitizer and drug delivery carrier can integrate photodynamic therapy (PDT) and chemotherapy. Via the strong coordination interaction between the zirconium cluster of porMOF and the terminal phosphate group of DNA, the stable modification of the DNA layer on the porMOF surface is achieved. Meanwhile, the introduction of C/G-rich base pairs into the DNA double-stranded structure provides more binding sites of chemotherapeutic drug doxorubicin (DOX). AS1411, an aptamer of nucleolin proteins that are overexpressed by cancer cells, is introduced in the double-stranded terminal, which can endow the nanosystem with the ability to selectively recognize cancer cells. C-rich sequences in DNA double strands form an i-motif structure under acidic conditions to promote the highly efficient release of DOX in cancer cells. In vitro and in vivo experiments demonstrate that the synergistic PDT/chemotherapy modality achieves highly efficient cancer cell killing and tumor ablation without undesirable side effects

Low-Background CRISPR/Cas12a Sensors for Versatile Live-Cell Biosensing

The trans-cleavage activity of CRISPR/Cas12a has been widely used in biosensing. However, many CRISPR/Cas12a-based biosensors, especially those that work in “on–off–on” mode, usually suffer from high background and thus impossible intracellular application. Herein, this problem is efficiently overcome by elaborately designing the activator strand (AS) of CRISPR/Cas12a using the “RESET” effect found by our group. The activation ability of the as-designed AS to CRISPR/Cas12a can be easily inhibited, thus assuring a low background for subsequent biosensing applications, which not only benefits the detection sensitivity improvement of CRISPR/Cas12a-based biosensors but also promotes their applications in live cells as well as makes it possible to design high-performance biosensors with greatly improved flexibility, thus achieving the analysis of a wide range of targets. As examples, by using different strategies such as strand displacement, strand cleavage, and aptamer–substrate interaction to reactivate the inhibited enzyme activity, several CRISPR/Cas12a-based biosensing systems are developed for the sensitive and specific detection of different targets, including nucleic acid (miR-21), biological small molecules (ATP), and enzymes (hOGG1), giving the detection limits of 0.96 pM, 8.6 μM, and 8.3 × 10–5 U/mL, respectively. Thanks to the low background, these biosensors are demonstrated to work well for the accurate imaging analysis of different biomolecules in live cells. Moreover, we also demonstrate that these sensing systems can be easily combined with lateral flow assay (LFA), thus holding great potential in point-of-care testing, especially in poorly equipped or nonlaboratory environments

pH-Controlled Intracellular in Situ Reversible Assembly of a Photothermal Agent for Smart Chemo-Photothermal Synergetic Therapy and ATP Imaging

To advance anti-tumor efficiency and lessen the adverse effect caused by nanodrug residues in the body, a smart nanoagent system is developed and successfully used in intracellular ATP imaging and in vivo chemo-photothermal synergetic therapy. The nanoagent system is facilely prepared using a DNA complex to modify gold nanoparticles (AuNPs). The DNA complex is formed by three oligonucleotides (ATP aptamer, rC-DNA, and rG-DNA). The CG-rich structure in a ternary DNA complex could be exploited for payload of chemotherapeutic medicine doxorubicin (DOX), thus making efficient DOX transport into the tumor site possible. In tumor cells, especially in acidic organelles (e.g., endosome and lysosome), DOX could be rapidly released via the dual stimuli of overexpressed ATP and pH. What is more, the specific recognition of a fluorescently labeled aptamer strand to ATP can achieve the intracellular ATP imaging. pH-controlled reversible folding and unfolding of intermolecular i-motif formed by C-rich strands can lead to intracellular in situ assembly of AuNP aggregates with high photothermal conversion efficiency and promote relatively facile renal clearance of AuNPs through the disassociation of the aggregates in extracellular environments. Experiments in vivo and vitro present feasibility for a synergetic chemo-photothermal therapy. Such an in situ reversible assembly strategy of a chemo-photothermal agent also presents a new paradigm for a smart and highly efficient disease treatment with reduced side effects

Chiral Interaction Is a Decisive Factor To Replace d‑DNA with l‑DNA Aptamers

Nucleic acid aptamers have been widely used in various fields such as biosensing, DNA chip, and medical diagnosis. However, the high susceptibility of nucleic acids to ubiquitous nucleases reduces the biostability of aptamers and limits their applications in biological contexts. Therefore, improving the biostability of aptamers becomes an urgent need. Herein, we present a simple strategy to resolve this problem by directly replacing the d-DNA-based aptamers with left-handed l-DNA. By testing several reported aptamers against respective targets, we found that our proposed strategy stood up well for nonchiral small molecule targets (e.g., Hemin and cationic porphyrin) and chiral targets whose interactions with aptamers are chirality-independent (e.g., ATP). We also found that the l-DNA aptamers were indeed endowed with greatly improved biostability due to the extraordinary resistance of l-DNA to nuclease digestion. With respect to other small-molecule targets whose interactions with aptamers are chirality-dependent (e.g., kanamycin) and biomacromolecules (e.g., tyrosine kinase-7), however, the proposed strategy was not entirely effective likely due to the participation of the DNA backbone chirality into the target recognition. In spite of this limitation, this strategy indeed paves an easy way to screen highly biostable aptamers important for the applications in many fields

A Rapid and Facile Detection for Specific Small-Sized Amino Acids Based on Target-Triggered Destruction of Metal Organic Frameworks

Most of the reported metal organic frameworks (MOFs)-based DNA sensors were developed by utilizing the different adsorption capacities of MOFs to different structural DNAs (for example, single-stranded DNAs (ssDNAs) and double-stranded DNAs (dsDNAs)) or ssDNAs with different lengths. Herein, we introduced another strategy for the design of MOFs-based biosensing platforms. We found that specific small-sized amino acids (for example, glycine and serine) could lead to the destruction of the MOFs formed by [Cu­(mal)­(bpy)]·2H₂O], thus recovering the fluorescence of a fluorophore-labeled ssDNA that had been quenched by MOFs. The corresponding working mechanism was discussed. On the basis of this finding, a mix-and-detect fluorescence method was designed for the turn-on detection of specific small-sized amino acids. The feasibility of its use in real serum samples was also demonstrated. Besides biosensing applications, the discovery of amino acids-triggered destruction of MOFs can also enrich the building blocks of molecular logic gate. As an example, a biomolecular logic gate that performs OR logic operation was constructed using glycine and a DNA strand as inputs

“RESET” Effect: Random Extending Sequences Enhance the Trans-Cleavage Activity of CRISPR/Cas12a

The trans-cleavage activity of CRISPR/Cas12a has been widely used in biosensing applications. However, the lack of exploration on the fundamental properties of CRISPR/Cas12a not only discourages further in-depth studies of the CRISPR/Cas12a system but also limits the design space of CRISPR/Cas12a-based applications. Herein, a “RESET” effect (random extending sequences enhance trans-cleavage activity) is discovered for the activation of CRISPR/Cas12a trans-cleavage activity. That is, a single-stranded DNA, which is too short to work as the activator, can efficiently activate CRISPR/Cas12a after being extended a random sequence from its 3′-end, even when the random sequence folds into secondary structures. The finding of the “RESET” effect enriches the CRISPR/Cas12a-based sensing strategies. Based on this effect, two CRISPR/Cas12a-based biosensors are designed for the sensitive and specific detection of two biologically important enzymes