110 results about "Conformational change" patented technology

Provided herein are products and methods for detecting analytes using polymers that bind to such analytes and thereby undergo a conformational change or contribute to a newly formed complex.

The invention discloses a deep learning prediction method fused with multi-modal features, which can accurately identify protein hidden binding sites in a ligand-free (apoo) state. The method comprises the following steps of: firstly, constructing a protein graph by taking residues as nodes and taking C alpha distance less than or equal to 14 as edges, wherein node feature sets comprise amino acid one-hot, secondary structures, atomic attributes, protein language model embedding and BLOSUM62 evolutionary information, and edge features comprise distance and angle similarity; then capturing three-dimensional geometric equivariant features by adopting an equivariant graph neural network (EGNN), and modeling a chemical topological relation by using a graph isomorphic network (GINE) with edge features; eGNN and GINE double-branch feature fusion and global dependence integration are realized through gating cross attention and gating multi-head attention; and finally, inputting the fusion features into a Kolmogorov-Arnold network (KAN) classifier, and predicting whether each residue belongs to a hidden binding site or not. The method can adapt to large-scale conformation change without coordinate alignment, AUC and F1 on a standard data set are remarkably superior to those of an existing method, high robustness and generalization are kept for multi-chain protein and complex conformation, and the method can be widely applied to drug target discovery and structure-driven drug design.

The invention relates to the technical field of gene editing, in particular to a Cas9 protein mutant and application thereof. According to the present invention, the mutation is introduced at the position far away from the catalytic site to induce the Cas9 protein conformation change so as to obtain the new Cas9 protein mutant, only the Cas9-sgRNA target chain is cut, and the Cas9-sgRNA non-target chain is not cut, such that the new Cas9 protein mutant can be used for constructing the novel base editor, and the high-efficiency base editing can be performed on the mammalian cell genome. The Cas9 protein mutant disclosed by the invention is fused with a mutant TX with DNA binding deletion of 3 '-> 5' exonuclease TREX2, so that the gene editing efficiency of a 3'terminal generated by paired single-nick enzyme cutting can be greatly improved, and the safety of the Cas9 protein mutant is maintained.

The invention discloses an influenza A virus neutralizing antibody and application thereof, the neutralizing antibody comprises a heavy chain and a light chain, the heavy chain comprises a heavy chain CDR1 with an amino acid sequence as shown in SEQ ID NO: 5, a heavy chain CDR2 with an amino acid sequence as shown in SEQ ID NO: 6 and a heavy chain CDR3 with an amino acid sequence as shown in SEQ ID NO: 7; the light chain comprises a light chain CDR1 with an amino acid sequence as shown in SEQ ID NO: 8, a light chain CDR2 with an amino acid sequence as shown in SEQ ID NO: 9 and a light chain CDR3 with an amino acid sequence as shown in SEQ ID NO: 10. The antibody disclosed by the invention has high-efficiency broad-spectrum neutralizing activity in influenza A virus, and can be combined with various subtypes of hemagglutinin and inhibit splitting of hemagglutinin precursor protein and conformational change of the hemagglutinin in an acid environment. The invention has important significance and application prospect for preventing and treating diseases caused by influenza A virus.

The invention provides a method for analyzing interaction between procambarus clarkia arginine kinase and shark source nano antibody. The method comprises the following steps: performing three-dimensional structure prediction on a VNAR sequence, and screening a high-confidence VNAR model by using a Laplace map; docking an amino acid sequence corresponding to the VNAR model with an arginine kinase sequence to obtain an optimal compound model, and analyzing an interaction site of an arginine kinase epitope region and a nanometer antibody complementary determining region CDR3; performing molecular dynamics simulation on the compound model, calculating conformational change indexes and binding free energy through energy optimization, system balance and extended simulation sampling, and outputting a binding stability sequence of the nano antibody and arginine kinase; the method comprises the following steps: screening out a high-stability nano antibody, pre-incubating a fusion protein of the high-stability nano antibody and immobilized arginine kinase to form a compound, adding serum of an allergic patient for competitive binding, quantifying the binding activity of residual IgE through an immunodetection technology, and verifying the inhibition effect of the shark source nano antibody on the sensitization effect of arginine kinase.

The invention relates to a modular polypeptide comprising a first partial effector sequence comprising a first part of a circular permutated halotag protein connected to a sensor module sequence, which is connected to a second part of a circular permutated halotag protein. The sensor module is a single polypeptide or a polypeptide pair capable of undergoing conformational change from a first confirmation to a second confirmation depending on the presence or concentration of an analyte compound. The modular peptide is catalytically active in response to an environmental stimulus or in response to the sensor pair interacting.The invention further relates to nucleic acid sequences encoding the modular polypeptide, and to kits comprising same.

The embodiment of the invention discloses a virus three-dimensional reconstruction method and system based on three-dimensional Gaussian splashing. The method comprises the following steps: acquiring three-dimensional point cloud data including a target virus; performing molecular dynamics simulation on the target virus based on a coarse graining molecular dynamics model, identifying a high-energy interaction region, and obtaining binding energy data of the target virus and a host molecule in an interaction process; tracing the conformation change of the virus structure to obtain conformation change data; performing three-dimensional Gaussian splashing processing on the point cloud data to generate an in-situ visual three-dimensional model of the target virus; wherein in the Gaussian splashing processing, the shape, the direction and the weight of a Gaussian kernel are dynamically adjusted according to combined energy data and conformation change data. According to the method and the device, efficient virus three-dimensional model generation is realized aiming at virus characteristics.

The invention discloses a preparation method and application of an electrochemical biosensor for dynamically monitoring Cyt c in tumor cells based on fast scanning voltammetry. The preparation method is characterized by comprising the following steps: 1) preparing a PtNE nano electrode; (2) preparing the PtNE / Apt nano electrochemical biosensor; and (3) accurately puncturing the PtNE / Apt electrode into cytoplasm of a single Hela cell through a three-dimensional micro-control system, so that Apt and intracellular Cyt c are specifically combined and subjected to conformational change, and 3 '-end modified Fc is close to the surface of the electrode, thereby generating a remarkable electrochemical signal. The change of Fc redox peak current is dynamically monitored through fast scanning voltammetry, the quantitative relation between the Cyt c concentration and the peak current is established, and the relation between the Cyt c concentration and the cell activity is monitored according to the quantitative relation. The device has the advantages of high sensitivity, excellent anti-interference capability and remarkable signal amplification effect.

Disclosed are a recombinant interferon drug targeting a conserved HIV gp41 subunit membrane-proximal external region (MPER) and a use thereof. Specifically, disclosed is a targeted interferon fusion protein ZP-IC, comprising a 10E8 antibody or an antigen-binding fragment thereof, and an integrated interferon α, wherein the 10E8 antibody or the antigen-binding fragment thereof is linked to the integrated interferon α by means of a linker. In the present invention, the 10E8 antibody capable of identifying a highly conserved gp41 subunit MPER of an HIV envelope glycoprotein is used as a targeting tool for the integrated interferon. Therefore, a target exposed by a conformational change of a viral envelope protein during the infection of a cell can be accurately identified, viral infection is prevented, and a neutralizing effect is exerted; and in addition, a carried interferon molecule can more effectively bind to an interferon receptor on a cell around the HIV virus and achieve an antiviral role. Experiments have demonstrated that the ZP-IC has excellent antiviral activity and the capability of inhibiting an overwhelming majority of HIV strains across a broad spectrum.

The invention provides a disease marker binding pocket identification method, and belongs to the technical field of disease marker binding pockets, and the method comprises the following steps: firstly, constructing a simulation system containing a disease marker, and carrying out 100-500ns molecular dynamics simulation; a conformation change track is collected and subjected to normalization processing; carrying out decomposition calculation on the sampled conformation, and analyzing a conformation cluster with relatively high frequency; calculating structural parameters of amino acid residues, and screening a region with relatively high flexibility as a potential binding site; analyzing the solvent accessibility and electrostatic characteristics of the sites; identifying a temporary binding pocket through the water molecule distribution density; performing principal component analysis to extract a conformation change mode; constructing an amino acid residue interaction network, and screening a stable binding pocket based on indexes such as binding free energy, conformational stability and pocket comprehensive performance evaluation; the method solves the problem that most existing methods only consider static structural characteristics of molecules and neglect the influence of dynamic change of protein conformation on binding pocket formation.

The invention discloses a non-competitive detection method based on aptamer allosteric activation CRISPR / Cas and application, and belongs to the technical field of biosensing. The detection method solves the problems that in a CRISPR / Cas detection system based on the nucleic acid aptamer in a competitive mode, the design requirement for partial complementary chains of the nucleic acid aptamer is high, and multiple requirements of low background signals, efficient target recognition and effective activation of the CRISPR / Cas system are difficult to consider. According to the detection method, the conformational change in the process that the aptamer recognizes the target is converted into the dynamic difference of the footrest-mediated strand displacement reaction, and the cascade footrest-mediated strand displacement reaction is designed to activate the CRISPR / Cas system. The detection method realizes high-sensitivity, simple, convenient and rapid ATP detection, and is applied to freshness evaluation of pork, beef and mutton.

Disclosed are systems, methods, and computer software for determining a conformational change in a structure of a protein. One method includes delivering a magnetic nanoparticle-nanobody (MNP-NB) complex to a sample containing a protein, where the MNP-NB complex will bind to the protein in the sample. An external magnetic field is applied to the sample with a magnetic field generation system. Signals are detected from the MNP-NB complex that reflect a response to the external magnetic field and a conformational change in a structure of the protein in the sample is determined from the signals.

According to the method for detecting the conductance of the protein molecules and the application of the method, asymmetric modification is carried out on the tunneling electrode pair and the protein molecules, so that the protein molecules can enter a tunneling area in a fixed posture and form asymmetric single-molecule junctions with the tunneling electrode pair, the activity of the protein molecules can be kept to the maximum extent, and the conductivity of the protein molecules is detected. Therefore, the device capable of stably detecting the conductance of the protein molecules for a long time can be constructed and obtained, and the device can be used for researching the dynamic conformation of the protein, screening small-molecule drugs targeting the protein molecules and analyzing the interaction relationship between the small-molecule drugs and the protein molecules. The tunneling electrode device and the real-time electrical measurement technology are used for achieving protein detection on the single molecule level, the conformation change process of protein such as biological enzyme and the action characteristics of the protein and small molecules are comprehensively described on the high time resolution scale, and the design and optimization efficiency and success rate of existing drugs are expected to be improved.

Methods of labeling, processing and analyzing protein samples. Surface-accessible residues in a protein sample (such as a proteome) can be labeled using a carbodiimide coupling agent an amine comprising a click chemistry handle, such as glycine propargyl amide. The click chemistry handle can then be derivatized using high-yield "click" chemistry to form a stable linkage with an enrichable label. The proteome can then be digested, peptides that have been modified can be enriched, and the enriched peptides can be analyzed, for example, by mass spectrometry. The analyzed peptides can constitute a highly enriched subset of the original peptides while still representing many sites of modification on many proteins (~95%) originally present in the proteome. The methods can be used to identify solvent-accessible regions of proteins, conformational changes, and drug targets across a whole proteome.

Disclosed are protein switches, components of such protein switches, and uses thereof, which can sequester biologically active peptides and / or binding domains, keeping them in an inactive ("off") state until combined with a second, designed polypeptide, termed a key, thereby inducing a conformational change that activates ("on") the biologically active peptide or binding domain.

The invention discloses a cell membrane in-situ drug membrane protein target screening method based on variable configuration DNA, and belongs to the technical field of biological medicine. Two pairs of functionalized nucleic acid structures are constructed, the first pair is S1S2 semi-complementary DNA double strands, the second pair is S3S4 DNA-RNA hybrid double strands, and the S1S2 structure is covalently coupled to a non-natural amino acid site-directed modified membrane protein through a click chemical reaction in a living cell to realize tagging of the membrane protein; when the candidate drug coupled in S3 interacts with the membrane protein, triggering an allosteric response system to release the tagged chain in S1; the released DNA tag is subjected to PCR amplification and sequencing, and accurate identification and analysis of a membrane protein target are realized according to a tag sequence. The invention provides a high-specificity and non-in-vitro drug membrane protein target screening technology. According to the technology, active compound discovery or accurate screening of drug membrane protein targets in a living cell in-situ environment can be realized.

The invention relates to an antibody-biomacromolecule conjugate. The antibody-biomacromolecule conjugate comprises an antibody targeting module, a complement effect module and a connexon, the antibody targeting module can be specifically combined with a cell membrane surface antigen, and the complement effect module is a key enzyme C1s in a classical complement pathway or a functional structural domain with complement splitting activity of the key enzyme C1s. The linker is used for connecting the antibody targeting module with the complement effect module, and is designed to generate cutting or conformational change on the outer side of a cell membrane or in an extracellular microenvironment, so that the complement effect module is converted from a limited state to a functional accessible state; to promote initiation of a classical complement pathway in the neighborhood of the cell membrane located by the antibody targeting module. The antibody-biomacromolecule conjugate provided by the invention can be used for engineering regulation and control of complement-related reactions.

The application discloses a VEGF / TIE-2 double-targeting polypeptide and application thereof in preparation of an antitumor drug, and relates to the technical field of biological medicine.The VEGF / TIE-2 double-targeting polypeptide (named as RTP) provided by the application is composed of an amino acid sequence capable of target-recognizing VEGF and TIE-2, an amino acid sequence capable of self-assembling to form a beta-sheet nanofiber, an amino acid sequence capable of target-recognizing carbonic anhydrase IX and an enzyme-reactive amino acid sequence capable of being specifically cut by MMP-2.RTP can occur conformational change and self-assemble in tumor cells to form stable water-insoluble nanofibers, and can strongly inhibit tumor neovascularization and metastasis through the VEGF and TIE-2 double-signal pathways and inhibit the progress of the tumor.

The present invention discloses a molecular dynamics simulation analysis method for ultrasonic perturbation of the Piezo1 protein, belonging to the field of ultrasonic perturbation. In the molecular dynamics simulation, ultrasonic perturbation is achieved by applying an ultrasonic function to the boundary, and successful ultrasonic loading is verified based on the water density on the X-axis. Furthermore, an analytical method is established to explain the effects of ultrasound on cell membranes from multiple angles, such as cell membrane thickness and lipid-protein interactions, and further study the effects of cell membranes on proteins, thereby determining the effects of ultrasound on the Piezo1 structure. This invention is the first to use ultrasonic loading coarse-grained molecular dynamics simulation to study the conformational changes of the Piezo1 mechanosensitive ion channel, providing important research ideas for future applications in ultrasound therapy, acoustic genetics, and other fields.

The present invention provides a method for detecting the electrical conductivity of protein molecules and its application. By asymmetrically modifying a tunneling electrode pair and a protein molecule, the protein molecule can enter the tunneling region in a fixed posture and form an asymmetric single-molecule junction with the tunneling electrode pair, which can maximize the activity of the protein molecule. This constructs a device that can stably detect the electrical conductivity of protein molecules over a long period of time. The device can be used for dynamic conformational studies of the protein itself, screening of small molecule drugs targeting the protein molecule, and analysis of the interaction between small molecule drugs and protein molecules. The present invention utilizes tunneling electrode devices and real-time electrical measurement technology to achieve protein detection at the single-molecule level, comprehensively depicting the conformational change process of proteins such as biological enzymes and their interaction characteristics with small molecules on a high time resolution scale, and is expected to improve the efficiency and success rate of existing drug design and optimization.

The invention discloses a neutralizing antibody with an AQP3 channel blocking function as well as a preparation method and application thereof, and belongs to the technical field of biological medicines. The neutralizing antibody can be specifically combined with an extracellular domain of AQP3, after combination, a water channel and a small molecule transport function of the AQP3 are blocked through steric hindrance and conformation change, and meanwhile, antigen combination specificity is reserved. The preparation method is realized through'recombinant antigen-immunization-B cell screening culture-recombinant antibody expression and affinity purification-ELISA detection ', and the functions of the antibody are ensured. The antibody can be efficiently combined with AQP3 positive cells, is strong in targeting property and high in specificity (the serum titer reaches 1: 1024 * 10, and the purified antibody titer is greater than or equal to 1: 128 * 10), and can be independently used or combined with a nano-carrier for treating related diseases, so that the technical bottleneck that a common AQP3 antibody can only be combined and cannot efficiently play a blocking role is solved.

The invention discloses a method for detecting the conformational change of an APO-Cascade protein. Through mutation of Cas8 protein amino acid N376 and Cas5 protein K169 in a Casade protein, the obtained Cas8 and Cas5 labeled by non-natural amino acids can maintain functions, and at the same time, the mutant type of the Cas5 / 8 protein can be subjected to fixed-point fluorescence labeling; according to the method, the Cas6 is used as a template, the N end of the Cas6 is further modified with biotin, and a formed Casade compound can be fixed on a biotin-PEG functionalized single-molecule slide, so that the conformation state of the Casade protein can be truly reflected by using single-molecule fluorescence resonance energy transfer, an effective means is provided for in-vitro research on the structure-function relationship of a system in CRISPR / Casade, and the method has a wide application prospect. And the method is of great significance in designing the Cascade protein with optimized properties and enabling the Cascade protein to play new functions.

The invention belongs to the technical field of medicine and pharmacy, and particularly relates to application of an AP1B1 allosteric activator of ethyl gallate. In the invention, AP1B1 is a beta 1 subunit (AP1B1) of a linking factor associated protein complex 1 (AP-1), and is an essential component for assembling a functional AP-1B complex. The ethyl gallate is used as an allosteric activator of AP1B1 and is specifically combined with an allosteric pocket of the AP1B1 to induce the AP1B1 to generate conformational change, so that the vesicle transportation function of an AP1B complex is activated, and the substrate-side membrane protein sorting capacity is recovered. A target dependence experiment verifies that the effect of ethyl gallate on promoting tight junction protein ZO-1 expression and membrane localization in AP1B1 knocked-down Caco-2 cells is remarkably weakened, and it is prompted that the intestinal barrier protection function of ethyl gallate depends on AP1B1. The invention provides a new strategy for developing medicines for treating intestinal barrier injury related diseases.

This invention discloses a neutralizing antibody against influenza A virus and its applications. The neutralizing antibody has a heavy chain and a light chain. The heavy chain comprises heavy chain CDR1 (amino acid sequence as shown in SEQ ID NO:5), heavy chain CDR2 (amino acid sequence as shown in SEQ ID NO:6), and heavy chain CDR3 (amino acid sequence as shown in SEQ ID NO:7); the light chain comprises light chain CDR1 (amino acid sequence as shown in SEQ ID NO:8), light chain CDR2 (amino acid sequence as shown in SEQ ID NO:9), and light chain CDR3 (amino acid sequence as shown in SEQ ID NO:10). The antibody of this invention exhibits highly efficient and broad-spectrum neutralizing activity against influenza A virus, and can bind to multiple subtypes of hemagglutinin, inhibiting the cleavage of hemagglutinin precursor proteins and conformational changes of hemagglutinin in acidic environments. This invention has significant implications and promising applications for the prevention and treatment of diseases caused by influenza A virus.

The invention provides a transition state searching method and system for complex biomacromolecule conformational change and application. The method comprises the steps that an input data set related to molecular dynamics simulation trajectory data is acquired; dimensionality reduction is carried out on an input data set through a reversible dimensionality reduction method, trajectory data is projected from a high-dimensional space to a low-dimensional space on the basis of reserving transition state information, and a low-dimensional free energy surface is generated; searching a low-dimensional candidate transition state sample on the low-dimensional free energy surface by adopting a low-dimensional search method; inversely projecting the low-dimensional candidate transition state sample back to a high-dimensional space through a reversible dimension reduction method to obtain a complete candidate transition state structure; and verifying the candidate transition state structure, and screening out a real transition state structure according to a verification result. According to the method, high-dimensional transition state structure search with high calculation efficiency and high accuracy is realized through reversible dimension reduction and low-dimensional search based on generative artificial intelligence, and the method is high in expandability, high in physical reliability and suitable for a complex biomolecular system.

Disclosed herein are sensor chips, imaging systems, and methods of preparing and using the same. The sensor chips disclosed herein include a substrate and a plurality of nanoplasmon rulers immobilized on the substrate. Each nanoplasmon ruler includes a first plasmonic element and a second plasmonic element connected by opposing ends of a single strand nucleic acid connector. Binding of an analyte to the single strand nucleic acid connector induces a conformational change in the single strand nucleic acid connector. The conformational change modulates an end-to-end distance between the first plasmonic element and the second plasmonic element, thereby modulating the nanoplasmonic coupling between the first plasmonic element and the second plasmonic element.

The invention discloses a cell single-molecule mechanical force sensor for single-molecule mechanical force detection in a cell phagocytosis process as well as a preparation method and application of the cell single-molecule mechanical force sensor. The sensor comprises a microsphere substrate and a plurality of double-stranded DNA mechanical probes modified on the surface of the microsphere substrate, the probe is composed of a first single-stranded DNA and a partially complementary second single-stranded DNA and forms an FRET or fluorescence quenching pair, when a pulling force applied to the probe by a receptor-ligand effect exceeds a preset melting threshold value in a process of swallowing or wrapping microspheres by a cell, the double-stranded DNA generates melting or conformational change and generates fluorescence signal change, and the fluorescence signal change is detected by the first single-stranded DNA and the partially complementary second single-stranded DNA. Therefore, the detection of the monomolecular mechanical force of the phagocytic interface is realized. By adjusting the geometric conformation and sequence design of the probe, a probe library with different melting force thresholds can be constructed, and quantitative analysis and spatial distribution characterization of mechanical force in the phagocytosis process are realized. According to the application, a three-dimensional phagocytic interface can be truly simulated, and an effective tool is provided for revealing a mechanism for regulating and controlling endothelial cell phagocytic behaviors through blood flow mechanics and screening related disease drugs.

The invention provides a method for detecting mycotoxin by using a carbon nanotube field effect transistor biosensor based on nucleic acid conformation change, which comprises the following steps: anchoring a modified nucleic acid aptamer on the surface of an ion shielding gold film layer through a gold-sulfur bond by using a CNT-FET biosensor with an ion shielding gold film layer above a channel to construct a sensing interface; the modified nucleic acid aptamer consists of a target mycotoxin original nucleic acid aptamer sequence and an oligomeric thymine deoxynucleotide fragment directionally inserted into the 5'end of the target mycotoxin original nucleic acid aptamer sequence; during detection, mycotoxin in a to-be-detected sample is specifically combined with the modified nucleic acid aptamer, secondary conformation change of the mycotoxin close to the gold film layer is triggered, the CNT-FET biosensor converts an electric field effect caused by conformation displacement into a channel carrier concentration change signal, and quantitative detection is achieved according to the linear relation between the signal and the mycotoxin concentration. According to the method, the nucleic acid aptamer is directionally modified, so that the electrical response after the low-charge-quantity / electrically neutral mycotoxin is combined with the sensor is enhanced, and the detection sensitivity is greatly improved.

The present invention discloses a porous silicon deoxyribozyme fluorescent probe and its application in evaluating the anticancer activity of traditional Chinese medicine. The fluorescent probe comprises amino-modified porous silica nanoparticles, fluorescent molecules loaded in the nanoparticle pores, and DNA hairpins HP1 and HP2 covalently linked to the nanoparticle surface. The HP1 sequence contains a complementary sequence to the microRNA to be detected, and the HP2 sequence contains a zinc ion-specific deoxyribozyme cleavage site. After HP1 binds to the microRNA, the root is opened, the HP1 loop undergoes a conformational change, and binds to the HP2 loop nucleotide sequence. With the assistance of zinc ions, the zinc ion-specific deoxyribozyme is activated, which then cuts HP2 at the cleavage site, releasing the fluorescent molecules in the nanoparticle pores. The microRNA is detected based on the intensity of the fluorescent molecules. The probe can specifically and sensitively detect trace amounts of microRNA in a system.

The present invention relates to methods, kits, and compositions for testing a sample from a subject and determining: i) if said subject is A / A, A / C, or C / C at position 4444 in the Glutamyl-prolyl-tRNA synthetase 1 (EPRS1) gene or mRNA, and / or ii) if said subject expresses only the 1482T version, only the 1482P version, or both the 1482T and 1482P versions, of the EPRS1 protein; and / or iii) if said subject is m6A methylated or m6A unmethylated at A4355 and / or A4464 in said EPRS1 mRNA; and determining that the subject has hypomyelinating leukodystrophy (HLD) or other neurological condition. The present application also relates to a bioinformatic pipeline to assess if m6A-distal single-nucleotide-variations (m6Ad-SNV) affect methylation of DRACH sites.