Precise Structural Analysis of Neutral Glycans Using Aerolysin Mutant T240R Nanopore.

Glycans play vital roles in nearly all life processes of multicellular organisms, and understanding these activities is inseparable from elucidating the biological significance of glycans. However, glycan research has lagged behind that of DNA and protein due to the challenges posed by structural heterogeneity and isomerism (i.e., structures with equal molecular weights) the lack of high-efficiency structural analysis techniques. Nanopore technology has emerged as a sensitive single-molecule biosensor, shining a light on glycan analysis. However, a significant number of glycans are small and uncharged, making it challenging to elicit identifiable nanopore signals. Here we introduce a R-binaphthyl tag into glycans, which enhances the cation-π interaction between the derivatized glycan molecules and the nanopore interface, enabling the detection of neutral glycans with an aerolysin nanopore. This approach allows for the distinction of di-, tri-, and tetrasaccharides with monosaccharide resolution and has the potential for group discrimination, the monitoring of enzymatic transglycosylation reactions. Notably, the aerolysin mutant T240R achieves unambiguous identification of six disaccharide isomers, trisaccharide and tetrasaccharide linkage isomers. Molecular docking simulations reveal that multiple noncovalent interactions occur between residues R282, K238, and R240 and the glycans and R-binaphthyl tag, significantly slowing down their translocation across the nanopore. Importantly, we provide a demonstration of the kinetic translocation process of neutral glycan isomers, establishing a solid theoretical foundation for glycan nanopore analysis. The development of our technology could promote the analysis of glycan structural isomers and has the potential for nanopore-based glycan structural determination and sequencing.

Precise Capture and Dynamic Release of Circulating Liver Cancer Cells with Dual-Histidine-Based Cell Imprinted Hydrogels.

Circulating tumor cells (CTCs) detection presents significant advantages in diagnosing liver cancer due to its noninvasiveness, real-time monitoring, and dynamic tracking. However, the clinical application of CTCs-based diagnosis is largely limited by the challenges of capturing low-abundance CTCs within a complex blood environment while ensuring them alive. Here, an ultrastrong ligand, l-histidine-l-histidine (HH), specifically targeting sialylated glycans on the surface of CTCs, is designed. Furthermore, HH is integrated into a cell-imprinted polymer, constructing a hydrogel with precise CTCs imprinting, high elasticity, satisfactory blood compatibility, and robust anti-interference capacities. These features endow the hydrogel with excellent capture efficiency (>95%) for CTCs in peripheral blood, as well as the ability to release CTCs controllably and alive. Clinical tests substantiate the accurate differentiation between liver cancer, cirrhosis, and healthy groups using this method. The remarkable diagnostic accuracy (94%), lossless release of CTCs, material reversibility, and cost-effectiveness ($6.68 per sample) make the HH-based hydrogel a potentially revolutionary technology for liver cancer diagnosis and single-cell analysis.

A highly sensitive nanochannel device for the detection of SUMO1 peptides.

SUMOylation is an important and highly dynamic post-translational modification (PTM) process of protein, and its disequilibrium may cause various diseases, such as cancers and neurodegenerative disorders. SUMO proteins must be accurately detected to understand disease states and develop effective drugs. Reliable antibodies against SUMO2/3 are commercially available; however, efficient detectors are yet to be developed for SUMO1, which has only 50% homology with SUMO2 and SUMO3. Here, using phage display technology, we identified two cyclic peptide (CP) sequences that could specifically bind to the terminal dodecapeptide sequence of SUMO1. Then we combined the CPs and polyethylene terephthalate conical nanochannel films to fabricate a nanochannel device highly sensitive towards the SUMO1 terminal peptide and protein; sensitivity was achieved by ensuring marked variations in both transmembrane ionic current and Faraday current. The satisfactory SUMO1-sensing ability of this device makes it a promising tool for the time-point monitoring of the SENP1 enzyme-catalyzed de-SUMOylation reaction and cellular imaging. This study not only solves the challenge of SUMO1 precise recognition that could promote SUMO1 proteomics analysis, but also demonstrates the good potential of the nanochannel device in monitoring of enzymes and discovery of effective drugs.

Nanofluidic Device for Detection of Lysine Methylpeptides and Sensing of Lysine Methylation.

Protein methylation is the smallest possible yet vitally important post-translational modification (PTM). This small and chemically inert addition in proteins makes the analysis of methylation more challenging, thus calling for an efficient tool for the sake of recognition and detection. Herein, we present a nanofluidic electric sensing device based on a functionalized nanochannel that was constructed by introducing monotriazole-containing p-sulfonatocalix[4]arene (TSC) into a single asymmetric polymeric nanochannel via click chemistry. The device can selectively detect lysine methylpeptides with subpicomole sensitivity, distinguish between different lysine methylation states, and monitor the lysine methylation process by methyltransferase at the peptide level in real time. The introduced TSC molecule, with its confined asymmetric configuration, presents the remarkable ability to selectively bind to lysine methylpeptides, which, coupled with the release of the complexed Cu ions, allows the device to transform the molecular-level recognition to the discernible change in ionic current of the nanofluidic electric device, thus enabling detection. This work could serve as a stepping stone to the development of a new methyltransferase assay and the chemical that specifically targets lysine methylation in PTM proteomics.

Geometrical and magnetic properties of small titanium and chromium clusters on monolayer hexagonal boron nitride.

Magnetic clusters on an insulating substrate are potential candidates for spin-based quantum devices. Here we investigate the geometric, electronic, and magnetic structures of small Ti and Cr clusters, from dimers to pentamers, adsorbed on a single-layer hexagonal boron nitride (h-BN) sheet within the framework of density functional theory. The stable adsorption configurations of the Ti clusters and Cr clusters composed of the same number of atoms are found to be totally different from each other. The difference in their bonding mechanisms has been revealed by the density of states and the charge density difference of the corresponding adsorption systems. While chemical bonds are formed between the Ti atoms and the supporting sheet, the Cr clusters are found in the physisorption state on the substrate. In addition, it is shown that the h-BN sheet is energetically favorable for building three-dimensional Ti clusters. These findings support the use of h-BN as a suitable decoupling substrate for manipulation of quantum spin states in small transition metal (TM) clusters and fabrication of devices based on them.

Spatially Resolved Stimulation for the Controlled Debromination in Single Molecules on a Surface.

A controlled chemical reaction on a specific bond in a single molecule is an inevitable step toward atomic engineering and fabrication. Here, we explored the debromination of a single 9,10-dibromoanthracene (DBA) molecule on a surface as stimulated by the voltage pulse through the tip of a scanning tunneling microscope (STM). A voltage threshold of about 2.2 V is obtained, and the nature of single-electron process is revealed. The spatially resolved debromination yield is obtained as a function of the pulse magnitude, which presents strong asymmetry for the two C-Br bonds. The optimal stimulation parameters including the pulse magnitude and the tip locations are suggested. The distinct dynamics in dissociation of the two bonds are illustrated by their energy diagrams and recoil paths, as derived by the first-principles density functional theory (DFT) calculation. The influence of the local electric field due to the STM tip on the dissociation of the C-Br bond has also been discussed. The study presents detailed practice for the controlled debromination in a single DBA molecule, which may lead to automated atomic engineering and fabrication of artificial nanostructures in the future.

Ionic Current Fluctuation and Orientation of Tetrahedral DNA Nanostructures in a Solid-State Nanopore.

Understanding the dynamic behavior of a nanostructure translocating through a nanopore is important for various applications. In this paper, the characteristics in ion current traces of tetrahedral DNA nanostructures (TDN) translocating through a solid-state nanopore are examined, by combined experimental and theoretical simulations. The results of finite element analysis reveal the correlation between orientation of TDN and the conductance blockade. The experimentally measured fluctuations in the conductance blockade, expressed as voltage-dependent histogram profiles, are consistent with the simulation, revealing the nature of a random distribution in orientation and weak influence of electrostatic and viscous torques. The step changes in orientation of a TDN during translocation are further explained by the collision with the nanopore, while the gradual changes in orientation illustrate the impact of a weak torque field in the nano-fluidic channel. The results demonstrate a general method and basic understanding in the dynamic behavior of nanostructures translocating through solid-state nanopores.

Detection and Characterization of Single Cisplatin Adducts on DNA by Nanopore Sequencing.

Detection and characterization of an individual cisplatin adduct on a single DNA molecule is a demanding task. We explore the characteristic features of cisplatin adducts in the nanopore sequencing signal in aspects of dwell time, genome anchored current trace, and basecalling accuracy. The offset between the motor protein and the nanopore constriction region is revealed by dwell time analysis to be about 14 bases in the nanopore device as we examined. Characteristic distortions due to cisplatin adducts are illustrated in genome anchored current trace analysis, constituting the fingerprint for identification of cisplatin adduct. The sharp increase in odds ratio at the location of adducting sites provides additional feature in the detection of the adduct. By these combined methods, single cisplatin adducts can be detected with high fidelity on a single read of the DNA sequence. The study demonstrates an effective method in the detection and characterization of single cisplatin adducts on DNA at the single-molecule level and with single nucleotide spatial resolution.

Revealing Differential Interaction Forces during Nanopore DNA Sequencing.

The interaction between DNA and the nanopore structure plays an important role in nanopore DNA sequencing. Differential interaction forces between each base type and the nanopore structure are obtained by examining the correlation between translocation dwell time and the sequence. The viscous drag force and the intermolecular interaction are identified with single-nucleotide resolution. Active hydrogen donors and acceptors on the inner wall of the nanopore structure are revealed at various offset coordinates. The differential forces as demonstrated in this study have great potential in probing active hydrogen bond interaction in a single protein molecule with subnanometer spatial resolution.

Development and evaluation of a color-image-based visual roughness measurement method with illumination robustness.

At present, the application of machine vision methods for roughness measurement in production sites is limited by its adaptability to illumination variations during the measurement. In this study, a machine vision method for roughness measurement with robustness to illumination is proposed so as to explore the functions of its color image indices in improving the mathematical expression of the vector of three primary colors. Besides, virtual images of different-roughness surfaces were analyzed, the effects of the samples' surface texture orientations on measurement indices were discussed, and the singular value ratio was derived as an index for evaluating roughness. The experimental results showed that the samples' index values remained unchanged when the illumination was increased for both vertical and horizontal surface textures, indicating that the proposed method has strong robustness to illumination. In addition, the experimental results were verified by a support vector machine (SVM)-based method using 10 different-roughness test samples, with the verification range of 0.127-2.245 µm. It was found that the measurement accuracy reached 90%, suggesting that the proposed method is reasonable and feasible, and shows certain potential to be applied in engineering.

Partition level multiview subspace clustering.

Multiview clustering has gained increasing attention recently due to its ability to deal with multiple sources (views) data and explore complementary information between different views. Among various methods, multiview subspace clustering methods provide encouraging performance. They mainly integrate the multiview information in the space where the data points lie. Hence, their performance may be deteriorated because of noises existing in each individual view or inconsistent between heterogeneous features. For multiview clustering, the basic premise is that there exists a shared partition among all views. Therefore, the natural space for multiview clustering should be all partitions. Orthogonal to existing methods, we propose to fuse multiview information in partition level following two intuitive assumptions: (i) each partition is a perturbation of the consensus clustering; (ii) the partition that is close to the consensus clustering should be assigned a large weight. Finally, we propose a unified multiview subspace clustering model which incorporates the graph learning from each view, the generation of basic partitions, and the fusion of consensus partition. These three components are seamlessly integrated and can be iteratively boosted by each other towards an overall optimal solution. Experiments on four benchmark datasets demonstrate the efficacy of our approach against the state-of-the-art techniques.

Translocation of tetrahedral DNA nanostructures through a solid-state nanopore.

Tetrahedral DNA nanostructures (TDNs) are programmable DNA nanostructures that have great potential in bio-sensing, cell imaging and therapeutic applications. In this study, we investigate the translocation behavior of individual TDNs through solid-state nanopores. Pronounced translocation signals for TDNs are observed that are sensitive to the size of the nanostructures. TDNs bound to linear DNA molecules produce an extra signal in the ionic current traces. Statistical analysis of its relative temporal position reveals distinct features between TDNs bound to the end and those bound to the middle of the linear DNA molecules. A featured current trace for two TDNs bound to the same linear DNA molecule has also been observed. Our study demonstrates the potential of using TDNs as sensitive bio-sensors to detect specific segments of a single DNA molecule in real time, based on solid-state nanopore devices.