27 results about "Membrane anchor" patented technology

The invention provides an anti-PD-1 antibody, a CAR-T cell, and a preparation method and application thereof. The anti-PD-1 antibody comprises LCDR-1-3 as shown in SEQ ID NO: 1-3 and HCDR-1-3 as shown in SEQ ID NO: 4-6, respectively. The CAR-T cell expresses a novel element for efficiently blocking the PD-1, and the element is a single-chain antibody for targeting the PD-1 in a cell membrane anchoring manner. And the chimeric antigen receptor and the cell membrane anchored anti-PD-1 scFv are connected by a 2A cleavage protein. The membrane anchor type anti-PD-1 scFv expressed by the CAR-T cell can almost completely block the expression of PD-1 on the surface of a T cell membrane, and further block a signal channel combined with PD-1 / PD-L1, so that the capability of T cell depletion caused by antagonism tumor of the CAR-T cell is enhanced, and the purpose of enhancing the anti-tumor effect of the CAR-T cell is achieved.

The invention relates to the field of biological materials, in particular to an amphiphilic block copolymer self-assembled biomimetic membrane and a molecular membrane anchoring method and application. Wherein the biomimetic membrane is formed by self-assembly of membrane forming molecules; the film-forming molecule comprises an amphiphilic triblock copolymer, the configuration of the amphiphilic triblock copolymer is a hydrophilic chain segment A-hydrophobic chain segment B-hydrophilic chain segment A, and the amphiphilic triblock copolymer is modified by an organic silicon surfactant and then is self-assembled into the amphiphilic triblock copolymer biomimetic film. And the biological nanopore protein is promoted to be embedded into the biomimetic membrane by adopting anchoring molecules through a molecular membrane anchoring method, so that the application of the biomimetic membrane in preparation of a biosensor is realized. An organic silicon surfactant is added as a bridge, polarity repulsion between the substrate and the polymer is eliminated, and the triblock polymer is stably attached to the surface of the substrate. Anchoring molecules are adopted to form covalent bonds with the surface of the membrane and biomacromolecules to realize anchoring, so that the stability is extremely high, the elution resistance and the environmental interference resistance are high, and the method is suitable for biosensors.

This invention discloses a target-gated ratiometric pH sensor based on a dual CRISPR system of tetrahedral framework nucleic acids, belonging to the fields of biomedicine and nucleic acid nanotechnology. The sensor uses tetrahedral framework nucleic acids as a rigid scaffold, precisely assembling Cas12a, Cas13a, and their corresponding fluorescent reporter molecules in a 1:1 stoichiometric ratio. Their orthogonal pH responses form the basis of the ratiometric sensing. Target recognition-pH sensing logic gating is achieved through target-specific crRNA design. Cholesterol modification allows for stable anchoring on the lipid membrane surface. This invention also discloses the sensor's fabrication, membrane anchoring, pH monitoring, and target-gated function implementation methods. This sensor possesses advantages such as high-precision ratio measurement, accurate spatial arrangement, high specificity, stable membrane anchoring, and a wide pH response range, and can be used for precise monitoring of pH dynamics at biomembrane interfaces and during extracellular vesicle fusion.

The invention discloses a PVDF (Polyvinylidene Fluoride) membrane material anchoring structure in a strong wind zone of a high-altitude dry-hot valley, and relates to the technical field of air membrane buildings. The PVDF membrane material anchoring structure comprises an anchoring foundation, a mounting seat, a screw rod and an anchoring plate. The anchoring foundation comprises a steel reinforcement framework and concrete filler. The concrete filler is poured into the steel reinforcement framework. And the mounting seat is embedded into the concrete filler and connected with the steel reinforcement framework. The mounting base has a first mounting plate. A first surface on one side of the first mounting plate in the thickness direction is flush with the surface of the concrete filler and is exposed out of the surface of the concrete filler. The anchor plate overlaps the first mounting plate. And the anchoring plate and the first mounting plate are used for clamping two sides of the edge part of the membrane ceiling. One end of the screw rod is perpendicular to the first surface, penetrates through the first mounting plate and is embedded into the concrete filler, and the other end of the screw rod sequentially penetrates through the membrane ceiling and the anchoring plate and is connected with the anchoring plate. According to the scheme, the first surface of the first mounting plate is exposed, so that the sealing performance of the air film building is improved.

The invention relates to an HER2 protein detection sequence group based on a tetrahedral nucleic acid framework and catalytic hairpin assembly, which comprises tetrahedral skeleton sequences S1, S2, S3 and S4, a cholesterol modified sequence Chol, an aptamer chain sequence L and hairpin probes H1, H2 and H3, and the 5'end of the tetrahedral skeleton sequence S1 extends to form an HER2 aptamer sequence HApt; the aptamer chain sequence L and the HER2 aptamer sequence HApt part are complementarily paired; the 3'ends of the tetrahedral skeleton sequences S2, S3 and S4 extend to form connecting sequences used for being combined with a cholesterol modification sequence Chol. By adopting the technical scheme, the invention provides the HER2 protein detection sequence group based on the tetrahedral nucleic acid framework and catalytic hairpin assembly, the kit and application thereof, and an HER2 in-situ detection technology with high stability, excellent membrane anchoring capability and signal amplification function.

The application discloses a Turn-Off type cell in-vivo tracing probe and a preparation method and application thereof, and the probe structure is composed of three parts, including a membrane anchoring part, an enzyme specificity response part and a fluorescent group. The probe can mark cells through the hydrophobic tail chain of the membrane anchoring part, at this time, the fluorescent group part emits near-infrared fluorescence, and shows a Turn state. When the adoptive cells are apoptotic in the body, the probe falls off from the cells. The free probe falling off is phagocytosed by macrophages in the body, in the intracellular macrophages, the enzyme specificity response part in the probe structure responds to enzyme bond breaking, the fluorescent group is dissociated, does not emit light, and shows an Off state. Therefore, the fluorescent probe can not only trace the in-vivo living cells and monitor the death and life of the cells in the body, but also can effectively overcome the fluorescent "pollution" phenomenon caused by the persistent existence of the fluorescent of the free probe falling off, and provides a new visual tool for monitoring the in-vivo fate of the living cell drugs.

This invention relates to a HER2 protein detection sequence set based on a tetrahedral nucleic acid framework and catalytic hairpin assembly, comprising tetrahedral backbone sequences S1, S2, S3, and S4, a cholesterol-modified sequence Chol, an aptamer chain sequence L, and hairpin probes H1, H2, and H3. The 5' end of the tetrahedral backbone sequence S1 extends to the HER2 aptamer sequence HApt; the aptamer chain sequence L is partially complementary to the HER2 aptamer sequence HApt; and the 3' ends of the tetrahedral backbone sequences S2, S3, and S4 extend to linker sequences for binding the cholesterol-modified sequence Chol. Using the above technical solution, this invention provides a HER2 protein detection sequence set, kit, and its application based on a tetrahedral nucleic acid framework and catalytic hairpin assembly, exhibiting high stability, excellent membrane anchoring ability, and signal amplification capabilities, thus providing an in-situ HER2 detection technology.

The invention discloses a targeted gating ratio type pH sensor of a double-CRISPR system based on tetrahedral framework nucleic acid, and belongs to the technical field of biomedicine and nucleic acid nanotechnology. According to the sensor, tetrahedral framework nucleic acid is used as a rigid support, Cas12a, Cas13a and corresponding fluorescent reporter molecules are accurately assembled according to the stoichiometric ratio of 1: 1, and orthogonal pH responses of the Cas12a and the Cas13a form a ratio type sensing basis; target recognition-pH sensing logic gating is realized through target specific crRNA (Complementary Ribonucleic Acid) design; after being modified by cholesterol, the compound can be stably anchored on the surface of a lipid membrane. The invention also discloses a preparation method of the sensor, a membrane anchoring method, a pH monitoring method and a target gating function realization method. The sensor has the advantages of high-precision ratio measurement, accurate spatial arrangement, high specificity, stable membrane anchoring, wide pH response range and the like, and can be used for accurately monitoring pH dynamics in a biological membrane interface microenvironment and extracellular vesicle fusion process.

The invention relates to a formate dehydrogenase mutant and application thereof, and belongs to the field of enzyme engineering and carbon dioxide reduction. The invention provides a formate dehydrogenase mutant. The formate dehydrogenase mutant comprises a formate dehydrogenase catalytic subunit FdhA, an electron transport subunit FdhB and a membrane anchoring protein subunit FdhC, compared with formate dehydrogenase, the formate dehydrogenase mutant has the advantages that a membrane anchor protein subunit FdhC is deleted, or the membrane anchor protein subunit FdhC and an electron transport subunit FdhB are deleted. Compared with the existing formate dehydrogenase, the formate dehydrogenase mutant provided by the invention has high activity and can tolerate oxygen at the same time. The formate dehydrogenase mutant provided by the invention shows huge potential value in the carbon dioxide immobilization industry.

The invention discloses a genetically engineered membrane-anchored neurotrophic factor, an exosome and a preparation method thereof, and relates to the technical field of biology. The invention provides a recombinant brain-derived neurotrophic factor protein. The amino acid sequence of the recombinant brain-derived neurotrophic factor protein is shown as SEQ ID NO: 1. The genetically engineered membrane anchored neurotrophic factor exosome provided by the invention comprises a recombinant BDNF, and the recombinant BDNF has a transmembrane structural domain, an extracellular structural domain and an exosome sorting domain, can be highly enriched on an exosome membrane, retains the physiological activity of the BDNF, and promotes nerve cell survival and synaptic formation.

The polypeptide membrane anchoring high-throughput screening method for GPCR is that the C terminal of GPCR is connected with tetracycline-controlled transcriptional activator (tTA) through tobacco etch virus protease substrate sequence (TEV-Substrate). When GPCR is combined with the membrane anchoring polypeptide, the TEV protease cuts the substrate to release tTA, activates the expression of luciferase reporter gene (Luc), and the cell expressing luciferase can detect the production amount of oxidized luciferin by using an enzyme label instrument, so that the accurate screening of the targeted polypeptide compound is realized. The screening method provided by the application can quickly screen the polypeptide combined with GPCR, is not limited to the screening of only a certain signal path in the traditional GPCR function detection, and can realize the high-throughput screening of the polypeptide and GPCR target by combining the luciferase reporter gene.

This invention discloses a recombinant yeast for producing α-ionone, its construction method, and its applications. The construction method involves using a yeast encoding osmanthus flowers... Osmanthus fragrans The gene for the carotenoid cleavage dioxygenase OfCCD1 was obtained through rational modification and codon optimization. OfCCD1m The gene, with its 5' end fused to the N-terminal membrane-anchored peptide coding sequence of human tyrosine kinase LCK, was transformed into the δ site of Saccharomyces cerevisiae TW3 to obtain recombinant yeast TW4 producing α-ionone. The Saccharomyces cerevisiae strain TW4 producing α-ionone of this invention has the following beneficial effects: it achieves de novo synthesis of α-ionone in Saccharomyces cerevisiae for the first time, filling a technological gap in this field; by using a membrane anchoring strategy to locate the OfCCD1m enzyme in the yeast plasma membrane, the contact efficiency between the enzyme and the substrate ε-carotene is significantly improved, resulting in an α-ionone yield of 1.46 mg / L.

The invention discloses gene modification carrying an immunological enhancement element and application of the gene modification, and relates to the technical field of biological medicine and gene engineering. B, at least one coding nucleic acid sequence IE of an immunological enhancement element, wherein the immunological enhancement element is used for enhancing antigen presentation, immunological co-stimulation signal transmission or positive regulation of a local immune microenvironment; c, at least one of a signal peptide segment SP and a membrane anchoring localization domain TM; the d expression regulation unit comprises a promoter Pr for driving the expression of Ag and IE and a transcription termination signal pA; the e-site strategy related sequence organization comprises a homologous segment left side HS-L and a homologous segment right side HS-R; wherein the Ag and the IE are assembled in the same expression framework, the IE is located at one of the upstream and the downstream of the Ag, antigen presentation and costimulatory signals are synchronous and stable, the immune response intensity and durability are improved, and inter-individual and inter-batch fluctuation is reduced.

The application discloses a kind of for preventing Bungowenia virus Polyepitope fusion protein, gene, vaccine and preparation method thereof, belong to biological medicine technical field.The Polyepitope fusion protein for preventing Bungowenia virus provided in the application is obtained in series by E. coli outer membrane anchoring protein ClyA, four MHC I class restricted T cell epitopes, four MHC II class restricted T cell epitopes, two B cell epitopes and BALB / c mouse immunoglobulin Fc fragment.The vaccine of the application is prepared by high-pressure homogenization process to obtain the engineered artificial nanovesicle (E-aBNV) with high-density surface display antigen array.The E-aBNV utilizes itself pathogen-associated molecular pattern to provide "self-adjuvant" effect, can efficiently activate B cell receptor and promote antigen cross-presentation, thereby synergistically inducing high level of neutralizing antibody and Th1-biased cellular immunity, which can effectively block the infection of Bungowenia virus to host cell.

This invention discloses a nucleic acid molecule encoding a modified retroviral Gag protein and its applications. The Gag protein contains an engineered acylated tag sequence at its N-terminus, comprising: a glycine residue at the second position after the initiating methionine for myristylation modification; and / or a cysteine ​​residue in the N-terminal region for palmitoylation modification. The tag sequence is selected from SEQ ID NO:1, 3-9, or derivative amino acid sequences having at least 80% identity and retaining function. Preferably, the modification involves replacing the first 9 amino acids at the N-terminus of the wild-type HIV-1 Gag protein with the aforementioned tag to enhance membrane anchoring and improve viral packaging efficiency. This invention also provides packaging plasmids containing this molecule, host cells, packaging systems, and preparation methods. This invention can significantly improve viral yield and titer, and is particularly suitable for overcoming the efficiency bottleneck in packaging large-load lentiviruses of 7-10 kb.

The application discloses a kind of knee joint targeting stem cell exosome compositions and preparation method and application thereof, it is related to biological medicine technical field.The composition includes the core-shell structure that is made of stem cell exosome and the targeting stable functional macromolecule that is coated on the surface of stem cell exosome;The targeting stable functional macromolecule is made of biocompatible polysaccharide skeleton, membrane anchoring hydrophobic group and knee joint lesion targeting ligand by covalent bond connection composition.The application is integrated in the same macromolecule carrier by integrated molecular design, and by self-assembly technology, it is " dressed " on the surface of exosome, solves the technical contradiction that stability and targeting cannot be obtained in existing stem cell exosome technology.

The present application provides a kind of CD47 mutant and its application, wherein CD47 mutant includes the amino acid sequence of partial amino acid sequence of CD47 and glycosyl phosphatidyl inositol (GPI) connection signal.The CD47 mutant is obtained by removing potential cell survival inhibition, blood flow and angiogenesis inhibition and other negative effect signal transduction sequences (transmembrane region and intracellular region) in CD47, while the beneficial functional segment (extracellular IgV domain) involved in inhibiting the phagocytosis of myeloid cells and its cytotoxic activity in CD47 sequence is fused with GPI membrane anchor connection signal sequence from human decay accelerating factor (DAF) protein.The obtained CD47 mutant can protect cells and organ transplants from being mediated by myeloid cells and other SIRP alpha positive immune cells with equal efficacy to wild type CD47, and at the same time eliminates the negative effects of wild type CD47.

The invention discloses injectable hemostatic hydrogel based on engineering modified cell derivatives as well as a preparation method and application of the injectable hemostatic hydrogel, and belongs to the technical field of biomedical materials. Chemical bonding and photo-crosslinking are cooperated, a membrane anchoring component is used for modifying the surface of a cell derivative to introduce an active chemical group, the active chemical group reacts with a hydrogel matrix complementary functional group to form a covalent bond, and a photoinitiator drives hydrogel to be crosslinked and cured under illumination, so that the cell derivative is stably fixed, burst release is avoided, and the biological function exerting efficiency is improved. The preparation method comprises the steps of engineering modification, stirring reaction, illumination crosslinking and the like, so that long-term stable loading of the cell derivative is realized. The hydrogel is high in biological safety and excellent in effect on substantive organ hemorrhage, and a new thought and scheme are provided for targeted delivery and controllable immobilization in the field of cell derivative-based biological materials.

Methods to purposefully target lytic granules to the cell membrane following NK cell activation for the benefit of bystander killing in a solid tumor setting as novel cell therapy approach. These methods can be used alone or in combination with dispersion related strategies. Methods of using truncated proteins from the dynein-dynactin complex to block dynein function to induce lytic granule dispersion. The methods and compositions enhance the lethality of degranulation events. The methods and compositions can boost cytotoxicity in a solid tumor directly or be combined with other technologies, such as immunotherapies to improve their outcomes.

The invention relates to a cell surface display system constructed based on sulfolobus membrane related protein and a construction method and application thereof, and belongs to the technical field of microbial genetic engineering. The sulfolobus pellicle-associated protein is a sulfolobus pellicle-associated protein SiRe2017, a sulfolobus pellicle-associated protein SiRe2108 or a sulfolobus pellicle-associated protein SiRe2467, and the amino acid sequences of the sulfolobus pellicle-associated protein SiRe2017, the sulfolobus pellicle-associated protein SiRe2108 or the sulfolobus pellicle-associated protein SiRe2467 are respectively shown The surface display system and the genetically engineered bacterium which take the Sulfolobus lellangii as a host are constructed by screening the membrane anchoring protein in the Sulfolobus lellangii, so that the field of host bacteria is expanded, and the application range of a microbial cell surface display technology, especially the application in an extreme environment, is widened. The linker between the Sulfolobus membrane associated protein SiRe anchoring motif and the reporter gene is also optimized, so that the Sulfolobus membrane associated protein SiRe anchoring motif is more suitable for a thermophilic and acidophilic environment, the activity and the display efficiency of the anchoring protein in an extreme environment are improved, and the activity of the anchoring protein is enhanced.

The invention discloses a GLP1 fusion protein with pancreas islet targeting, an exosome modified by the GLP1 fusion protein as well as a preparation method and application of the GLP1 fusion protein. The structure of the fusion protein is TP-linker-Lamp2b-linker-GLP1, the amino acid sequence of the pancreas islet targeting polypeptide TP is CPKTRRVY, and the pancreas islet targeting polypeptide TP can be specifically combined with pancreas islet beta cells; the amino acid sequence of the Linker is GSGSGSGSGS, so that the immunogenicity can be reduced, and the structural shielding can be avoided; the amino acid sequence of Lamp2b is as shown in SEQ ID NO.1, and exosome membrane anchoring can be realized; the amino acid sequence of the GLP1 mutant is HGEGTFPSDVSYLEGQAAKEFI AWLVKGR, and the half-life period of the GLP1 mutant is remarkably prolonged through A2G / T7P mutation. The invention also provides a preparation method of the fusion protein modified exosome. The preparation method comprises the steps of lentiviral vector construction, hUC-MSCs transduction and exosome extraction. In-vitro and in-vivo experiments prove that the modified exosome can target pancreas islet beta cells, efficiently exert the hypoglycemic effect of the GLP1 and remarkably improve the pancreas islet function, and a novel strategy which is high in targeting property, long-acting and efficient in delivery is provided for diabetes treatment.

Provided are a chimeric protein and a use thereof. The chimeric protein comprises: a transmembrane transport region, a target protein display region, a membrane anchoring region, and an intracellular signal transduction region which are sequentially connected, wherein the transmembrane transport region comprises a transmembrane transport peptide segment; the target protein display region comprises a target protein peptide segment; the membrane anchoring region comprises a membrane anchoring peptide segment; and the intracellular signal transduction region comprises a budding aid peptide segment.

The application discloses an engineered exosome medicine for targeted degradation of TGF-beta, a preparation method and application, and the medicine comprises: a TGF-beta Trap protein which is used for competitively combining with TGF-beta; a membrane anchoring protein which is used for embedding the TGF-beta Trap protein and anchoring the TGF-beta Trap protein to the surface of an exosome membrane; and a targeted biological recognition molecule which is connected to the exosome through protein fusion or click chemistry reaction and is used for targeting the engineered exosome medicine to tumor cells. The engineered exosome medicine provided by the application is a medicine which can effectively block and remove TGF-beta, can effectively weaken systemic toxicity and off-target side effects, and avoids problems such as complexity of exosome in-vitro modification, batch difference, drug activity initiation and the like.

Disclosed are devices, systems, and methods of manufacture and use of modular functionalized cellular nanoparticles that can bind a wide range of ligands, payloads, and functional substances onto a nanoparticle surface. In various embodiments, a cell membrane coating on the nanoparticle is engineered to express a membrane anchor that can readily form a covalent bond with any functional moiety modified with an appropriate peptide tag sequence.

The invention discloses a gene modification method of a fusion cytokine expression element and an immunotherapy application of the fusion cytokine expression element, and relates to the technical field of gene engineering and tumor immunotherapy. Comprising the following steps that A, a preset site is selected in a genome of a host cell, a first construct AL-B-C-B-AR is provided, AL is a first homologous arm located on the upstream of the preset site, AR is a second homologous arm located on the downstream of the preset site, B is an I-SceI incision enzyme recognition site, and C is a selection marker capable of being screened in the forward direction; inserting the B-C-B sequence into the predetermined loci by homologous directional integration to form a replaceable landing frame; wherein the preset site is preferably a safe port site and a non-essential intron segment of a target functional gene, the length of the homologous arm is preferably one hundred to one thousand bases, and after integration, the correctness of the 5'end and the 3 'end is confirmed through PCR breakpoint amplification and sequencing at the same time. The whole body exposure and toxicity are reduced.

The invention discloses a gene engineering modified CD20 targeting microvesicle / exosome as well as a preparation method and application thereof, and belongs to the technical field of biological nano-carrier preparation and targeting delivery. Aiming at the problems of poor targeting property, insufficient modification stability, high immune side reaction risk and the like of an existing extracellular vesicle (EV) delivery system, a CD20 antibody single-chain variable region (scFv) structural domain is fused with EV membrane anchoring structural domains such as Lamp2b, CD63, C1-C2, PDGFR beta and the like to construct a multi-type surface display carrier, so that stable presentation of the CD20 scFv structural domain on an EV membrane is realized. The engineered exosome / vesicle can specifically recognize CD20 positive B cells, improves the delivery efficiency, does not contain an Fc structure, avoids complement activation and related immunotoxicity, keeps the structural integrity of an EV natural membrane, and has large-scale production potential. The invention can be widely applied to the fields of treatment of CD20 positive B cell diseases, drug screening and transformational medicine, and has important clinical value.

Disclosed herein a robust approach to monitor cell interactions with its surrounding environment in real-time in a physiologically relevant 3-D microenvironment, holding great promise for enhancing future studies of detecting and monitoring cell processes and responses to instructive cues and therapeutic agents. A microfluidic cell culture system is provided that involves a cell sensor in a 3D culture scaffold and one or more microfluidic channels fluidly connected to the 3D culture matrix. The cell sensor involves a planar nucleic acid nanostructure having a top surface and a bottom surface, a plurality sensor molecule attached to the top surface to detect at least two target molecules, and one or more membrane anchoring moieties attached to the bottom surface.