ABPP-CoDEL: Activity-Based Proteome Profiling-Guided Discovery of Tyrosine-Targeting Covalent Inhibitors from DNA-Encoded Libraries.

DNA-encoded chemical library (DEL) has been extensively used for lead compound discovery for decades in academia and industry. Incorporating an electrophile warhead into DNA-encoded compounds recently permitted the discovery of covalent ligands that selectively react with a particular cysteine residue. However, noncysteine residues remain underexplored as modification sites of covalent DELs. Herein, we report the design and utility of tyrosine-targeting DELs of 67 million compounds. Proteome-wide reactivity analysis of tyrosine-reactive sulfonyl fluoride (SF) covalent probes suggested three enzymes (phosphoglycerate mutase 1, glutathione s-transferase 1, and dipeptidyl peptidase 3) as models of tyrosine-targetable proteins. Enrichment with SF-functionalized DELs led to the identification of a series of tyrosine-targeting covalent inhibitors of the model enzymes. In-depth mechanistic investigation revealed their novel modes of action and reactive ligand-accessible hotspots of the enzymes. Our strategy of combining activity-based proteome profiling and covalent DEL enrichment (ABPP-CoDEL), which generated selective covalent binders against a variety of target proteins, illustrates the potential use of this methodology in further covalent drug discovery.

Drug Development in the Field of Sphinogolipid Metabolism.

Sphingolipids are the major lipid components on cellular membranes especially on lipid raft regions, intermediating various important biological functions for eukaryotic cells. Sphingolipid metabolism pathways can utilize sugar, protein, nucleic acid, and other metabolites participating lipid transport in the circulation, play an essential role in maintaining cell homeostasis and are related to a variety of different diseases including lysosomal storage disorders (LSDs), Gaucher disease, etc. The dynamic balance of sphingolipid levels in organisms is regulated by a series of sphingolipid synthases, hydrolases, and metabolic enzymes, such as sphingomyelinase (SMase), sphingomyelin synthase (SMS), serine palmitoyltransferase (SPT), ceramide synthase (CerS), glucosylceramide synthase (GCS), etc. Thus, sphingolipids and its related enzymes are potential targets for drug discoveries and receive great research interests by medicinal chemist. In this chapter, we will discuss the relationship between sphingolipids and the regulating enzymes involved in sphingolipid metabolisms, and systematically summarize the advances in the development of new drugs in the field.

Directing Multivalent Aptamer-Receptor Binding on the Cell Surface with Programmable Atom-Like Nanoparticles.

Multivalent interactions of biomolecules play pivotal roles in physiological and pathological settings. Whereas the directionality of the interactions is crucial, the state-of-the-art synthetic multivalent ligand-receptor systems generally lack programmable approaches for orthogonal directionality. Here, we report the design of programmable atom-like nanoparticles (aptPANs) to direct multivalent aptamer-receptor binding on the cell interface. The positions of the aptamer motifs can be prescribed on tetrahedral DNA frameworks to realize atom-like orthogonal valence and direction, enabling the construction of multivalent molecules with fixed aptamer copy numbers but different directionality. These directional-yet-flexible aptPAN molecules exhibit the adaptability to the receptor distribution on cell surfaces. We demonstrate the high-affinity tumor cell binding with a linear aptPAN oligomer (≈13-fold improved compared to free aptamers), which leads to ≈50 % suppression of cell growth.

Prescribing Silver Chirality with DNA Origami.

Metal nanostructures of chiral geometry interacting with light via surface plasmon resonances can produce tailorable optical activity with their structural alterations. However, bottom-up fabrication of arbitrary chiral metal nanostructures with precise size and morphology remains a synthetic challenge. Here we develop a DNA origami-enabled aqueous solution metallization strategy to prescribe the chirality of silver nanostructures in three dimensions. We find that diamine silver(I) complexes coordinate with the bases of prescribed single-stranded protruding clustered DNA (pcDNA) on DNA origami via synergetic interactions including coordination, hydrogen bonds, and ion-π interaction, which induce site-specific pcDNA condensation and local enrichment of silver precursors that lowers the activation energy for nucleation. Using tubular DNA origami-based metallization, we obtain helical silver patterns up to a micrometer in length with well-defined chirality and pitches. We further demonstrate tailorable plasmonic optical activity of metallized chiral silver nanostructures. This method opens new pathways to synthesize programmable inorganic materials with arbitrary morphology and chirality.

Tailoring Oxygen-Containing Groups on Graphene for Ratiometric Electrochemical Measurements of Ascorbic Acid in Living Subacute Parkinson's Disease Mouse Brains.

Ascorbic acid (AA), a major antioxidant in the central nervous system (CNS), is involved in withstanding oxidative stress that plays a significant role in the pathogenesis of Parkinson's disease (PD). Exploring the AA disturbance in the process of PD is of great value in understanding the molecular mechanism of PD. Herein, by virtue of a carbon fiber electrode (CFE) as a matric electrode, a three-step electrochemical process for tailoring oxygen-containing groups on graphene was well designed: potentiostatic deposition was carried out to fabricate graphene oxide on CFE, electrochemical reduction that assisted in removing the epoxy groups accelerated the electron transfer kinetics of AA oxidation, and electrochemical oxidation that increased the content of the carbonyl group (C═O) generated an inner-reference signal. The mechanism was solidified by ab initio calculations by comparing AA absorption on defected models of graphene functionalized with different oxygen groups including carboxyl, hydroxyl, epoxy, and carbonyl. It was found that epoxy groups would hinder the physical absorption of AA onto graphene, while other functional groups would be beneficial to it. Biocompatible polyethylenedioxythiophene (PEDOT) was further rationally assembled to improve the antifouling property of graphene. As a result, a new platform for ratiometric electrochemical measurements of AA with high sensitivity, excellent selectivity, and reproducibility was established. In vivo determination of AA levels in different regions of living mouse brains by the proposed method demonstrated that AA decreased remarkably in the hippocampus and cortex of a subacute PD mouse than those of a normal mouse.

Programming nanoparticle valence bonds with single-stranded DNA encoders.

Nature has evolved strategies to encode information within a single biopolymer to program biomolecular interactions with characteristic stoichiometry, orthogonality and reconfigurability. Nevertheless, synthetic approaches for programming molecular reactions or assembly generally rely on the use of multiple polymer chains (for example, patchy particles). Here we demonstrate a method for patterning colloidal gold nanoparticles with valence bond analogues using single-stranded DNA encoders containing polyadenine (polyA). By programming the order, length and sequence of each encoder with alternating polyA/non-polyA domains, we synthesize programmable atom-like nanoparticles (PANs) with n-valence that can be used to assemble a spectrum of low-coordination colloidal molecules with different composition, size, chirality and linearity. Moreover, by exploiting the reconfigurability of PANs, we demonstrate dynamic colloidal bond-breaking and bond-formation reactions, structural rearrangement and even the implementation of Boolean logic operations. This approach may be useful for generating responsive functional materials for distinct technological applications.

Probing Transient DNA Conformation Changes with an Intercalative Fluorescent Excimer.

Variation of DNA conformation is important in regulating gene expression and mediating drug-DNA interactions. However, directly probing transient DNA conformation changes is challenging owing to the dynamic nature of this process. We show a label-free fluorescence method to monitor transient DNA conformation changes in DNA structures with various lengths and shapes using a DNA intercalator, K21. K21 can form transient excimers on the surface of DNA; the ratiometric emission of monomer and excimer correlate to DNA transient conformation stability in numerous DNA structures, including i-motifs, G-quadruplex structures, and single nucleotide mutation at random position. We analyzed the conformation dynamics of a single plasmid before and after enzyme digestion with confocal fluorescence microscopy. This method provides a label-free fluorescence strategy to probe transient conformation changes of DNA structures and has potential in uncovering transient genomic processes in living cells.

DNA Framework-Engineered Long-Range Electrostatic Interactions for DNA Hybridization Reactions.

Long-range electrostatic interactions beyond biomolecular interaction interfaces have not been extensively studied due to the limitation in engineering electric double layers in physiological fluids. Here we find that long-range electrostatic interactions play an essential role in kinetic modulation of DNA hybridizations. Protein and gold nanoparticles with different charges are encapsulated in tetrahedral frameworks to exert diverse electrostatic effects on site-specifically tethered single DNA strands. Using this strategy, we have successfully modulated the hybridization kinetics in both bulk solution and single molecule level. Experimental and theoretical studies reveal that long-range Coulomb interactions are the key factor for hybridization rates. This work validates the important role of long-range electrostatic forces in nucleic acid-biomacromolecule complexes, which may encourage new strategies of gene regulation, antisense therapy, and nucleic acid detection.

Poly-Adenine-Based Spherical Nucleic Acids for Efficient Live-Cell MicroRNA Capture.

Direct delivery of exogenous non-coding nucleic acids into living cells has attracted intense interest in biological applications. However, the cell entry efficiency and target capture ability remain to be improved. Herein, we report a method for compartmenting the nucleic acids on the surface of poly-adenine-based spherical nucleic acids (polyA-SNAs) for efficient capture of oncogenic microRNAs (miRNAs) in living cells. We find that polyA-SNAs exhibit high cell entry efficiency, which is insensitive to the configuration of the anti-miRNA sequences. By programming the length of polyAs, we precisely engineered the spatial configuration of the anti-miRNA sequences in polyA-SNAs. Compartmentalized polyA-SNAs bind to miRNAs with improved capture ability as compared to densely compacted SNAs. We further demonstrate that polyA-SNAs serve as high-efficacy miRNA sponges for capturing oncogenic miRNAs both in living cells and in mice. The efficient inhibition of miRNAs results in significant suppression of tumor growth.

Diverse Nanoassemblies of Graphene Quantum Dots and Their Mineralogical Counterparts.

Complex structures from nanoparticles are found in rocks, soils, and sea sediments but the mechanisms of their formation are poorly understood, which causes controversial conclusions about their genesis. Here we show that graphene quantum dots (GQDs) can assemble into complex structures driven by coordination interactions with metal ions commonly present in environment and serve a special role in Earth's history, such as Fe3+ and Al3+ . GQDs self-assemble into mesoscale chains, sheets, supraparticles, nanoshells, and nanostars. Specific assembly patterns are determined by the effective symmetry of the GQDs when forming the coordination assemblies with the metal ions. As such, maximization of the electronic delocalization of π-orbitals of GQDs with Fe3+ leads to GQD-Fe-GQD units with D2 symmetry, dipolar bonding potential, and linear assemblies. Taking advantage of high electron microscopy contrast of carbonaceous nanostructures in respect to ceramic background, the mineralogical counterparts of GQD assemblies are found in mineraloid shungite. These findings provide insight into nanoparticle dynamics during the rock formation that can lead to mineralized structures of unexpectedly high complexity.

The Marriage of Protein and Lanthanide: Unveiling a Time-Resolved Fluorescence Sensor Array Regulated by pH toward High-Throughput Assay of Metal Ions in Biofluids.

A protein/lanthanide complex (BSA/Tb3+)-based sensor array in two different pH buffers has been designed for high-throughput recognition and time-resolved fluorescence (TRF) detection of metal ions in biofluids. BSA, which acted as an antenna ligand, can sensitize the fluorescence of Tb3+ (i.e., antenna effect), while the presence of metal ions would lead to the corresponding conformational change of BSA for altering the antenna effect accompanied by a substantial TRF performance of Tb3+. This principle has also been fully proved by both experimental characterizations and coarse-grained molecular dynamics (CG-MD) studies. By using Tris-HCl buffer with different pHs (at 7.4 and 8.5), 17 metal ions have been well-distinguished by using our proposed BSA/Tb3+ sensor array. Moreover, the sensor array has the potential to discriminate different concentrations of the same metal ions and a mixture of metal ions. Remarkably, the detection of metal ions in biofluids can be realized by utilizing the presented sensor array, verifying its practical applications. The platform avoids the synthesis of multiplex sensing receptors, providing a new method for the construction of convenient and feasible lanthanide complex-based TRF sensing arrays.

Structural and positional impact on DNAzyme-based electrochemical sensors for metal ions.

The rapid, accurate and convenient detection of heavy metal is very important to public health. Here, we developed a DNAzyme-based electrochemical sensor for Pb2+. A DNAzyme-including and Pb2+ active probe was anchored to the biosensing interface, based on the well-defined self-assembled, three-dimensional DNA nanostructure. The results indicate that the detection performance depends on the change of distances between the methylene blue and the electrode surface. The limit of detection (LOD) could reach the concentration of 0.01 µM Pb2+, and the signal change shows semi-logarithmic relationship with the concentration of Pb2+ from 0.01 µM to 100 µM. The biosensor also presents good stability and specificity to detect Pb2+ in tap or river water. This method not only provides promising approach for improving the performance of tetrahedra in detecting Pb2+, but helps deepen the understanding of tetrahedral structure design and how the position of electroactive groups affects the performance of electrochemical sensing.

Chiral Ceramic Nanoparticles and Peptide Catalysis.

The chirality of nanoparticles (NPs) and their assemblies has been investigated predominantly for noble metals and II-VI semiconductors. However, ceramic NPs represent the majority of nanoscale materials in nature. The robustness and other innate properties of ceramics offer technological opportunities in catalysis, biomedical sciences, and optics. Here we report the preparation of chiral ceramic NPs, as represented by tungsten oxide hydrate, WO3-x·H2O, dispersed in ethanol. The chirality of the metal oxide core, with an average size of ca. 1.6 nm, is imparted by proline (Pro) and aspartic acid (Asp) ligands via bio-to-nano chirality transfer. The amino acids are attached to the NP surface through C-O-W linkages formed from dissociated carboxyl groups and through amino groups weakly coordinated to the NP surface. Surprisingly, the dominant circular dichroism bands for NPs coated by Pro and Asp are different despite the similarity in the geometry of the NPs; they are positioned at 400-700 nm and 500-1100 nm for Pro- and Asp-modified NPs, respectively. The differences in the spectral positions of the main chiroptical band for the two types of NPs are associated with the molecular binding of the two amino acids to the NP surface; Asp has one additional C-O-W linkage compared to Pro, resulting in stronger distortion of the inorganic crystal lattice and greater intensity of CD bands associated with the chirality of the inorganic core. The chirality of WO3-x·H2O atomic structure is confirmed by atomistic molecular dynamics simulations. The proximity of the amino acids to the mineral surface is associated with the catalytic abilities of WO3-x·H2O NPs. We found that NPs facilitate formation of peptide bonds, leading to Asp-Asp and Asp-Pro dipeptides. The chiroptical activity, chemical reactivity, and biocompatibility of tungsten oxide create a unique combination of properties relevant to chiral optics, chemical technologies, and biomedicine.

A single-wavelength-emitting ratiometric probe based on phototriggered fluorescence switching of graphene quantum dots.

Ratiometric fluorescent probes are of great importance in research, because a built-in correction for environmental effects can be provided to reduce background interference. However, the traditional ratiometric fluorescent probes require two luminescent materials with different emission bands. Herein a novel ratiometric probe based on a single-wavelength-emitting material is reported. The probe works by regulating the luminescent property of graphene quantum dots with UV illumination as activator. The ratiometric sensor shows high sensitivity and specificity for iron ions. Moreover, the ratiometric sensor was successfully employed to monitor ferritin levels in Sprague Dawley rats with chemical-induced acute liver damage. The proposed single-wavelength ratiometric fluorescent probe may greatly broaden the applicability of ratiometric sensors in diagnostic devices, medical applications, and analytical chemistry.

Hybrid nanotube-graphene junctions: spin degeneracy breaking and tunable electronic structure.

Hybrid carbon nanostructures have attracted enormous interest due to their structural stability and unique physical properties. Geometric and physical properties of a carbon nanotube (CNT)-graphene nanoribbon (GNR) hybrid system were investigated via first-principles density functional theory (DFT) calculations. The nanotube-graphene junction (NTGJ), where the GNR directly links to the CNT by covalent bonds, shows novel electronic dependence on the structural parameters of the building-blocks, such as chirality, nanotube diameter and width of the nanoribbon. For an armchair NTGJ, a small band gap opens up representing asymmetrical spin-up and spin-down bands. However, zig-zag NTGJ shows direct semi-conducting characteristics with a tunable band gap ranging from zero to 0.6 eV. Interestingly, the value of the band gap follows the specific width and diameter dependent oscillations, namely the 3p - 1 principle. Transition-state results reveal the formation of NTGJs is exothermic and has a low energy-barrier. In addition, nanotube-graphene-nanotube junctions or namely dumbbell NTGJs were also studied, which exhibits similar properties with single NTGJ.

Biomimetic Surface Engineering to Modulate the Coffee-Ring Effect for Amyloid-ß Detection in Rat Brains.

Surface engineering of nanoparticles has been widely used in biosensing and assays, where sensitivity was mainly limited by plasmonic colour change or electrochemical responses. Here, we report a novel biomimetic sensing strategy involving protein-modified gold nanoparticles (AuNPs), where the modulation strategy was inspired by gastropods in inhibition of coffee-ring effects in their trail-followings. The so-called coffee-ring effect presents the molecular behaviour of AuNPs to a macroscopic ring through aggregation, and thus greatly improves sensitivity. The assay relies upon the different assembly patterns of AuNPs against analytes, resulting in the formation or suppression of coffee-ring effects by the different surface engineering of AuNPs by proteins and peptides. The mechanism of the coffee-ring formation process is examined through experimental characterizations and computational simulations. A practical coffee-ring effect assay is developed for a proof-of-concept target, amyloid ß (1-42), which is a typical biomarker of Alzheimer's disease. A novel quasi-titrimetric protocol is constructed for quantitative determination of the target molecule. The assay shows excellent selectivity and sensitivity for the amyloid ß monomer, with a low detection limit of 20 pM. Combined with a fluorescent staining technique, the assay is designed as a smart sensor for amyloid ß detection and fibrillation evaluation in rat cerebrospinal fluids, which is a potential point-of-care test for Alzheimer's disease. Connections between amyloid fibrillation and different courses of brain ischaemia are also studied, with improved sensitivity, lower sample volumes that are required, convenience for rapid detection, and point-of-care testing.

High-entropy alloy nanopatterns by prescribed metallization of DNA origami templates.

High-entropy multimetallic nanopatterns with controlled morphology, composition and uniformity hold great potential for developing nanoelectronics, nanophotonics and catalysis. Nevertheless, the lack of general methods for patterning multiple metals poses a limit. Here, we develop a DNA origami-based metallization reaction system to prescribe multimetallic nanopatterns with peroxidase-like activities. We find that strong coordination between metal elements and DNA bases enables the accumulation of metal ions on protruding clustered DNA (pcDNA) that are prescribed on DNA origami. As a result of the condensation of pcDNA, these sites can serve as nucleation site for metal plating. We have synthesized multimetallic nanopatterns composed of up to five metal elements (Co, Pd, Pt, Ag and Ni), and obtained insights on elemental uniformity control at the nanoscale. This method provides an alternative pathway to construct a library of multimetallic nanopatterns.

Benzo[d]isoxazole Derivatives as Hypoxia-Inducible Factor (HIF)-1α Inhibitors.

Hypoxia-inducible factor, also known as HIF, is a transcriptional factor universally found in mammalian cells. HIF-1 is one of the HIF-families and acts as a heterodimer consisting of α and ß subunits. It is found to play significant roles in pathologic conditions such as tumor development and metastasis. Here, we first report benzo[d]isoxazole analogues as HIF-1α transcription inhibitors. Thereby, we designed and synthesized 26 benzo[d]isoxazole derivatives and evaluated their inhibitory activities against HIF-1α transcription in HEK293T cells by a dual-luciferase gene reporter assay. Among them, compounds 15 and 31 showed the best efficacy in a cell-based assay with an IC50 value of 24 nM and have potential antitumor effects for further development.

Mechanism of diastereoisomer-induced chirality of BiOBr.

Chiral molecule-driven asymmetric structures are known to be elusive because of the intriguing chirality transfer from chiral molecules to achiral species. Here, we found that the chiral assembly of BiOBr is independent of the chirality of the organic molecular inducer but dependent on geometric structural matching between the inducer and inorganic species. Diastereoisomeric sugar alcohols (DSAs) with identical numbers of carbon chiral centers and functional groups but with different R/S configurations and optical activities (OAs) were chosen as symmetry-breaking agents for inducing chiral mesostructured BiOBr films (CMBFs) under hydrothermal conditions. Multiple levels of chirality with different handedness were identified in the CMBFs. Density functional theory (DFT) calculations and molecular dynamics (MD) simulations suggest that asymmetric defects in the Br-Bi tetragonal cone caused by physically adsorbed DSAs on the surfaces of the BiOBr crystals are the geometric basis for triggering the chiral twist in the BiOBr monolayer. Our findings provide new insights for understanding the origin of chirality and the chiral transfer mechanism underlying the assembly of achiral species.

Modular Assembly of MXene Frameworks for Noninvasive Disease Diagnosis via Urinary Volatiles.

Volatile organic compounds (VOCs) in urine are valuable biomarkers for noninvasive disease diagnosis. Herein, a facile coordination-driven modular assembly strategy is used for developing a library of gas-sensing materials based on porous MXene frameworks (MFs). Taking advantage of modules with diverse composition and tunable structure, our MFs-based library can provide more choices to satisfy gas-sensing demands. Meanwhile, the laser-induced graphene interdigital electrodes array and microchamber are laser-engraved for the assembly of a microchamber-hosted MF (MHMF) e-nose. Our MHMF e-nose possesses high-discriminative pattern recognition for simultaneous sensing and distinguishing of complex VOCs. Furthermore, with the MHMF e-nose being a plug-and-play module, a point-of-care testing (POCT) platform is modularly assembled for wireless and real-time monitoring of urinary volatiles from clinical samples. By virtue of machine learning, our POCT platform achieves noninvasive diagnosis of multiple diseases with a high accuracy of 91.7%, providing a favorable opportunity for early disease diagnosis, disease course monitoring, and relevant research.