50 results about "Molecular modification" patented technology

This invention provides an in vitro cell-free protein synthesis system, its reagent kit, and its applications, belonging to the field of protein synthesis technology. This invention improves an in vitro cell-free protein synthesis system based on Kluyveromyces lactis by adding exogenous L-arabinose, significantly enhancing the system's protein synthesis capacity. Furthermore, it provides a more efficient and higher-throughput in vitro protein synthesis reagent kit and a method for synthesizing exogenous proteins. The improvements provided by this invention can be achieved without molecular modification, are simple and convenient to operate, and save costs.

The invention discloses a 3-sterone-delta1-dehydrogenase mutant and application of the 3-sterone-delta1-dehydrogenase mutant in preparation of a steroid drug intermediate. The mutant is obtained by performing single mutation or multiple mutation on 551, 468, 48 and 157 sites of an amino acid sequence of 3-sterone-delta1-dehydrogenase as shown in SEQ ID NO: 1. The 3-ketosterone-delta1-dehydrogenase is subjected to molecular modification through directed evolution and semi-rational design, the mutant with improved enzyme activity is screened in a high-throughput mode, the mutant can effectively improve the conversion rate of a substrate 11alpha, 17alpha-dihydroxyprogesterone, and the substrate conversion rate of the mutant ISM-2 is improved to 92.4% compared with 36.7% before mutation.

The invention discloses a surface-enhanced Raman probe as well as a preparation method and application thereof, and belongs to the technical field of biomedical imaging and stem cell therapy. The preparation method comprises the following steps: coating an ultrathin silver shell layer on the surface of gold nanoparticles to obtain Au-coated Ag particles; 4-sulfydryl phenylboronic acid pinacol ester molecules are modified on the surface of Au (at) Ag to form a Raman report layer, then sulfydryl-polyethylene glycol is introduced to realize biological orthogonal coupling, and the modified SERS probe is obtained through bovine serum albumin modification; jointly incubating the modified SERS probe and the azidation-labeled mouse bone marrow mesenchymal stem cells, so as to form an mmol / LSC-SERS composite probe; the obtained probe has good biocompatibility, stability and anti-interference capability and is suitable for a complex physiological environment, and the preparation method is simple and good in repeatability, is completely carried out in a water phase and does not need an organic solvent.

A method for producing a food product with gluten molecule modification, comprising the steps of: a) mixing at least gluten-containing flour with at least one liquid until a homogeneous mixture is obtained; b) containing the mixture in a gas-tight container and extracting air from the gas-tight container within 15 minutes after obtaining the mixture according to step a) in order to obtain a vacuum of at least equal to 70% inside the gas-tight container and maintaining the homogeneous mixture inside the container at a vacuum for a period of at least equal to 3 hours, thereby obtaining a mixture; c) obtaining a food product using the mixture obtained at the end of step b).

The application belongs to the field of bioengineering. The application provides a molecular modification design method for improving the hydrophobicity of the catalytic center of bacillus subtilis protease, which comprises the following steps: searching for a resolved bacillus subtilis protease crystal structure in a protein database PDB, generating an enzyme-substrate complex by docking a hexapeptide substrate through a Glide mode of software, calculating the polar charge of each amino acid residue of polar atoms N and O and H combined thereof within a 10 angstrom range of the catalytic center amino acid of the bacillus subtilis protease by using a PPC force field, calculating the absolute value sum of the polar charge, selecting a potential mutation site of the bacillus subtilis protease, virtually mutating the potential mutation site into a large steric hindrance nonpolar amino acid, calculating the absolute value sum of the polar charge of polar atoms N, O and H within a 10 angstrom range of the catalytic center amino acid of the single-point mutant by using the PPC force field, selecting a virtual bacillus subtilis protease single-point mutant, expressing the mutant and iteratively mutating, re-expressing, obtaining a modified bacillus subtilis protease mutant, and verifying the hydrophobicity. The molecular modification design method can quickly and effectively reduce water molecules in the catalytic center of the bacillus subtilis protease and improve the hydrophobicity thereof. It has been verified that the enzyme modification method is effective and reliable.

The present application relates to diaminopimelic acid dehydrogenase mutant and its application in D-phenylglycine synthesis. The present application discloses a diaminopimelic acid dehydrogenase mutant which is obtained by molecular modification of diaminopimelic acid dehydrogenase from Bacillus thermozeamaize, a nucleic acid coding the diaminopimelic acid dehydrogenase mutant, a recombinant expression vector containing the nucleic acid, a recombinant expression transformant containing the recombinant expression vector, and the use of the diaminopimelic acid dehydrogenase mutant in the synthesis of D-phenylglycine from asymmetric reduction of aminated benzoylformic acid. Compared with other D-phenylglycine synthesis methods, the route of the present application is simple, the theoretical yield is high, the atom economy is good, the substrate concentration and the product yield are high, and the optical purity of the product is greater than 99%. Therefore, the present application belongs to an environmentally friendly, high stereoselectivity and green and environmentally friendly biosynthesis method, and has good application prospect in the actual production of D-phenylglycine.

The application discloses a preparation method of chitosan microspheres for cell immobilization and drug delivery, and relates to the field of chitosan microsphere preparation. The application carries out deacetylation on chitin through a microwave induction process technology combined with enzymes, carries out heavy metal green chelation on chitosan by using glutamic acid diacetic acid tetrasodium or citric acid, further improves the extraction and removal capacity of the chelating agent on residual metal ions by using a carbonate salt solubilization technology, and separates and hydrolyzes trace amounts of protein and lipids in the crude chitosan by selecting appropriate biological enzymes to obtain refined chitosan. Meanwhile, the refined chitosan is subjected to enzymatic molecular modification by applying transglycosylation or oxidase, and finally, the chitosan microspheres are prepared by using a green and environment-friendly spray drying method. The chitosan microspheres prepared by the application are smooth in shape, uniform in size, have a particle size of 1-250 mu m, and have a shell-core structure. The chitosan immobilized enzyme widens the application of chitosan in the fields of medicine, food and the like.

This invention discloses an internal standard-assisted petal-shaped SERS nanoprobe, its preparation method, and its applications. The internal standard-assisted petal-shaped SERS nanoprobe of this invention is a core-molecule-shell structure gold nanoprobe consisting of a gold nanosphere core, an internal standard modifying molecule, a petal-shaped gold nanoshell, and a recognition molecule, 4-nitrobenzenethiophenol. The internal standard modifying molecule is a Raman signal molecule with nitrile, alkyne, or azide functional groups. This invention uses a Raman signal molecule as an internal standard modifying molecule to modify the gold nanosphere core. The corrected working curve exhibits good reproducibility and small error, improving the reliability of the detection results. Growing a petal-shaped gold nanoshell on the molecular layer of the internal standard modifying molecule enhances the SERS activity both inside and outside the SERS substrate, and the petal-shaped shell increases the surface area of ​​the SERS substrate. Furthermore, the recognition molecule of this invention enables ratiometric Raman detection of hydrogen sulfide, effectively reducing background signal interference and improving detection sensitivity.

The application provides a kind of tea polyphenol ester-RNA self-assembly and hybrid nanovesicle of probiotic outer membrane vesicle and preparation method thereof, which first adjusts the hydrophilicity and hydrophobicity of tea polyphenol EGC through molecular modification strategy, and the EGC palmitate obtained by modification can directly wrap RNA and self-assemble into tea polyphenol ester-RNA nanovesicle, which has superior encapsulation capacity. Further introduce OMVs as intestinal targeted delivery carrier and fuse with RNA-tea polyphenol nanovesicle to form a structure stable hybrid vesicle. The hybrid vesicle of the application ingeniously integrates the advantages of both sides: tea polyphenol ester-RNA nanovesicle ensures efficient encapsulation of RNA, and OMVs provide natural protection for its translocation through gastrointestinal tract, and promote intestinal cell uptake with its inherent bioadhesion. This strategy can effectively promote the encapsulation and stabilization of oral RNA delivery.

The invention relates to SERS (Surface Enhanced Raman Scattering) sensing test paper for detecting trace target molecules in tears as well as a preparation method and a detection method and belongs to the field of biosensing and spectral analysis. At least one of the problems of insufficient detection sensitivity, poor anti-pollution capability, low selectivity, incapability of rapid and on-site instant detection and the like in the existing tear biomarker detection technology is solved. The SERS sensing test paper comprises a porous fiber matrix and an SERS active nano composite material loaded on the porous fiber matrix, the SERS active nano composite material is precious metal nano particles (at) HZIF modified by Raman reporter molecules, and the SERS active nano composite material is of a core-shell structure. According to the invention, ultrasensitive, anti-pollution and rapid on-site detection of trace target molecules (such as histamine) in tears is realized.

The present application provides an anti-ultraviolet polydimethylsiloxane food packaging coating and a preparation method thereof, which realizes the dual synergy of anti-ultraviolet and food preservation function through molecular modification, and introduces degradable materials to enhance environmental protection. The coating embeds anti-ultraviolet groups and preservation groups into the polydimethylsiloxane main chain through chemical bonds, avoiding the migration of traditional additives. A one-step in-situ polymerization process is used to ensure uniform distribution of functional groups, solving the problems of uneven function and migration. The coating provides ultraviolet protection, antibacterial, ethylene absorption, water retention, and prolongs the shelf life of food. The coating also uses degradable materials to reduce environmental pollution. It is suitable for outdoor circulation and transparent packaging scenarios, providing an efficient, safe and environmentally friendly packaging solution.

The invention belongs to the technical field of biochemistry, and particularly discloses a novel LNP gene drug siRNA delivery carrier and a construction method thereof. The construction method comprises the following steps: by taking a DNA nanocube as a basic carrier, loading siRNA on the surface of the DNA cube in a base complementary pairing manner, then coating liposome on the outer layer of the cube by utilizing a DNA structure-assisted phospholipid membrane self-assembly technology, and modifying polypeptide target molecules on the outer surface of the liposome by adopting a post-insertion method, so as to obtain the siRNA-modified liposome. The siRNA nano delivery carrier capable of delivering siRNA to tumor tissues in a targeted manner is obtained. The delivery carrier constructed by the invention can realize targeted efficient delivery of siRNA, and effectively solves the following problems existing in current siRNA delivery: 1) the siRNA is easy to degrade in the delivery process, lysosome escape is difficult, and the delivery efficiency is low; 2) problems of potential cytotoxicity, easy induction of immune response and difficult metabolism of a delivery carrier, and 3) problems of liposome instability, low drug loading capacity and difficult size control.

The invention provides a modification method of an antibody, the modification method adopts a modification molecule with a cage-shaped structure to modify the antibody, and after modification, the rigid structure of the antibody molecule is enhanced, the stability is improved, and the half-life period is prolonged. Meanwhile, by adjusting the addition amount of the modified molecules, the obtained antibody can present a nanoparticle aggregation structure, and the activity of the antibody is further improved.

This invention discloses a bifunctional small-molecule PEG-modified recombinant humanized collagen and its preparation method. The method includes the following steps: preparing a borate buffer solution containing sodium cyanoborohydride, pH 7.5-8.5; dissolving lyophilized recombinant collagen powder in the buffer solution to obtain a substrate solution; adding crosslinking agent powder to the substrate solution according to a molar ratio of free lysine to crosslinking agent of 1:(1.5-4.0); the crosslinking agent is a bifunctional small-molecule PEG with PEG as the backbone and groups at both ends that can couple with the amino groups of collagen; reacting at 4°C in the dark for 2-5 hours. This invention not only overcomes the bottleneck of anti-enzymatic stability but also transforms recombinant humanized collagen into a multi-arm macromolecular crosslinking substrate with highly active groups on its surface, providing key chemical bonding sites for its subsequent targeted coupling or anchoring to polymer networks such as hydrogels.

The invention relates to a mannose isomerase mutant with improved enzyme activity and application of the mannose isomerase mutant. The amino acid sequence of the mannose isomerase mutant is shown as SEQ ID NO.4; the preparation method comprises the following steps: carrying out molecular modification on a Senotrophomonas maltophilia-sourced mannose isomerase, carrying out site-specific mutagenesis on proline at the 10th site of the Senotrophomonas maltophilia-sourced mannose isomerase to obtain alanine, carrying out site-specific mutagenesis on methionine at the 139th site of the Senotrophomonas maltophilia-sourced mannose isomerase to obtain isoleucine to obtain a mutant P10A + M139I, and purifying the mutant P10A + M139I through a nickel affinity chromatography column to obtain a pure enzyme; the optimal pH is 8.5, the optimal temperature is 60 DEG C, the relative enzyme activity is increased by 28.2% compared with that of a wild type, an excellent catalyst is provided for mannose production in the industry, the production cost of mannose is reduced, and a foundation is laid for research of mannose isomerase and mannose.

The invention relates to the technical field of chromatographic separation media, in particular to molecular size exclusion separation filler, which is prepared from the following raw materials in parts by weight: 40 to 80 parts of porous polysaccharide carrier; 5 to 20 parts of hydrophilic modification molecules; 5 to 15 parts of a coupling reagent; 5-10 parts of an alkaline regulator; 0.2 to 0.5 part of a stabilizing agent; and 16-20 parts of a dispersing aid. According to the invention, hydrophilic macromolecules are adopted to modify the surface of the pore channel, so that the size of the pore channel is effectively regulated and reduced, and a micromolecule sample has longer retention time. Compared with the method for reducing the pore diameter by increasing the use amount of raw materials, the method has the advantages that the characteristic that macromolecules with different molecular weights have different partition coefficients in pore channels is utilized, namely, the macromolecules can selectively enter different pore diameters relatively, namely, the molecules with small molecular weights modify the small pore diameters, and the molecules with large molecular weights modify the large pore diameters, so that the pore diameters are more gradient. The process method has the characteristics of lower cost and simpler operation.

The present application belongs to the technical field of biotechnology and enzyme engineering, and relates to a molecular modification method for improving substrate specificity of aldehyde ketone reductase, which is achieved by constructing a substrate library of aldehyde ketone reductase AKR13B3 and analyzing kinetic parameters, determining a mutation site by performing loop structure analysis and amino acid sequence alignment on a complex structure of aldehyde ketone reductase AKR13B3, coenzyme NADPH and substrate 3-keto-DON, and screening a mutant with increased substrate specificity, the present application designs the geometric shape of the entrance loop region of the substrate binding pocket of AKR13B3 to change substrate preference and realize the increase of catalytic activity of the enzyme on specific substrates, in addition, the present application provides a feasible scheme for designing aldehyde ketone reductase with good substrate specificity, and provides great hope for developing high-efficiency enzyme preparations.

Methods and kits for probing chromatin accessibility, including Deaminase Assisted single-molecule chromatin Fiber sequencing (DAF-seq) methods and kits. The disclosed approaches leverage DNA deaminase toxin A (DddA) to efficiently stencil protein occupancy along DNA molecules via selective deamination of sterically accessible cytosine bases. These deamination events result in C-to-T transitions during DNA amplification, such that the resultant sequences preserve the chromatin architectures of the underlying individual chromatin fibers as per-molecule DNA alterations. The disclosed approaches can be used for identifying where proteins are bound to genomic DNA, mappable at a single-cell, single-molecule, and single-nucleotide level, and as diagnostic tools in clinical settings.

The invention discloses a method for producing a clopidogrel intermediate through biological catalysis chiral resolution, which comprises the following steps: screening an esterase mutant capable of efficiently resolving the clopidogrel intermediate (R, S)-2-(2-chlorphenyl)-2-(2-thiophene ethylamino) methyl acetate through molecular modification to obtain (S)-2-(2-chlorphenyl)-2-(2-thiophene ethylamino) methyl acetate; according to the present invention, the biocatalysis activity and the stereoselectivity are high, the reaction efficiency is improved, the position of the biocatalysis step in the whole synthesis route is beneficial to the improvement of the efficiency of other chemical steps, the overall conversion rate, the enantiomer excess value and the product purity, and the production cost can be significantly reduced. In addition, the method also has the common advantages of enzymatic synthesis, such as simplicity and convenience in implementation and operation, greenness and environmental protection, easiness in product separation and the like.

The application provides a rational design method for improving thermal stability of lipase, and belongs to the field of enzyme engineering. A flexible region of the lipase to be reformed is determined through molecular dynamics simulation, a reforming region is determined through structure analysis, a reforming site is determined through comprehensive measurement of multiple calculation tools, a mutant library is established through site-directed saturation mutation using NNK degenerate codon, and an excellent mutant with improved thermal stability is screened through a microplate screening method based on spectrophotometry. The half-life of the Candida antarctica lipase B mutant of the application at 50 DEG C is 9 times that of the wild type, and the specific activity is also 6.32% higher than that of the wild type. The application breaks the trade-off between stability and activity, and realizes the improvement of the thermal stability of the lipase, which is conducive to the important role of the lipase mutant in the industrial application in the fields of food, medicine or biodiesel.

The invention discloses a method for improving the reliability of a phase delay film and the phase delay film, and belongs to the technical field of optics. According to the invention, the sulfur-sulfur alkyl chain-containing compound is adopted to perform molecular modification on the phase delay film, so that the adsorption and invasion rate of the phase delay film to water molecules can be reduced, and polymer swelling, plasticizing and hydrolysis effects caused by water vapor can be reduced; thermal motion of a polymer molecular chain in a high-humidity and high-temperature environment is inhibited, internal stress release and microstructure change are reduced, and then the problems of phase delay value drift, haze rising and surface physical degradation of the phase delay film in the high-humidity and high-temperature environment are solved; and the optical stability and the mechanical stability of the phase delay film in a high-humidity and high-temperature environment are improved.

The invention belongs to the technical field of beverage manufacturing, and discloses a preparation method of a Chinese wolfberry fruit beverage capable of improving hyaluronic acid absorbability through electron irradiation molecule modification, and the preparation method comprises the following steps: soaking Chinese wolfberry fruits, crushing, homogenizing and emulsifying to obtain a Chinese wolfberry fruit liquid; performing primary filtration, medium filtration and final filtration on the Chinese wolfberry liquid to obtain a small-molecular-grade Chinese wolfberry stock solution; adding the sodium hyaluronate solution into the micromolecular wolfberry stock solution according to the proportion of 0.2-0.5% to obtain a compound solution; and carrying out cold sterilization and small molecularization on the compound liquid by adopting an electron irradiation mode to obtain the Chinese wolfberry fruit beverage. According to the preparation method, cold sterilization and small molecularization are achieved, meanwhile, the absorption rate of sodium hyaluronate and fructus lycii nutrient substances is increased, the taste of the fructus lycii beverage is guaranteed, and nutritional ingredients in the raw materials are prevented from being damaged.

The invention discloses a modified carotenoid cleavage dioxygenase 1 and application thereof, and belongs to the technical field of protein engineering. According to the invention, molecular modification is carried out on the carotenoid cleavage dioxygenase 1 from petunia, site-directed mutagenesis and saturated mutation are carried out, screening is carried out to obtain a mutant of the carotenoid cleavage dioxygenase 1 with improved activity, the yield of beta-ionone generated by catalyzing the cleavage of beta-carotene by the mutant M142A / N174A / H244L / F306M is 2.85 times of that of wild PhCCD1, and the yield of beta-ionone generated by catalyzing the cleavage of beta-carotene is 2.85 times of that of the wild PhCCD1. The catalytic efficiency (kcat / Km) is improved by 1.7 times, and the mutant obviously improves the catalytic activity and catalytic efficiency of the carotenoid splitting dioxygenase 1 and lays a foundation for industrial application of the carotenoid splitting dioxygenase 1.

The invention provides a property-changed dihydroxyacetone kinase mutant and application thereof, and relates to the technical field of enzyme engineering. Through rational design and molecular modification, wild dihydroxyacetone kinase is subjected to single-point mutation and combined mutation, a series of enzyme mutants with enhanced thermal stability are obtained, and the catalytic activity of the mutants under the high-temperature condition is remarkably superior to that of wild enzymes. The invention further provides a multienzyme coupling reaction system containing the dihydroxyacetone kinase mutant, and the multienzyme coupling reaction system is used for efficiently catalyzing the process of synthesizing lactic acid by taking dihydroxyacetone as a substrate. The thermostable dihydroxyacetone kinase mutant provided by the invention solves the key bottleneck that a wild enzyme is easy to thermally inactivate under industrial application conditions, and has important industrial application prospects.

The invention belongs to the technical field of microorganisms, and particularly relates to a genetic transformation method aiming at Huiarenaceae microflora and application of the genetic transformation method. Specifically, a genetic transformation method aiming at Huiarenaceae microbe is established for the first time, exogenous plasmids are subjected to in-vitro methylation modification by using methylase in a cell-free system of the strain, and the activity of endonuclease is inhibited in combination with a combination mode of ATP consumption and EDTA chelation; the problem of exogenous plasmid degradation caused by a complex and unknown strain limitation modification system is effectively solved, and the immune barrier is successfully broken through. Meanwhile, by optimizing competent cell preparation conditions and electrotransformation parameters, the transformation efficiency reaches an operable level, and the requirements of molecular operation are met. The invention provides a key technical support for molecular modification of Huiaromatic Zhongaceae microflora, is helpful for excavating the metabolic potential of the Huiaromatic Zhongaceae microflora, improves the stress resistance, and promotes the industrial application of the Huiaromatic Zhongaceae microflora in the field of environmental governance such as combined pollution wastewater treatment.

The invention discloses an arteannuyl sphingosine base derivative as well as a preparation method and application thereof. The arteannuyl sphingosine base derivative provided by the invention is prepared by carrying out molecular modification on an arteannuic acid compound and a sphingosine base compound and carrying out amidation reaction. The derivative combines the biological activities of the two, enhances skin affinity, stability and functionality, has excellent multiple activities of resisting inflammation, relieving, improving skin immune barrier, resisting wrinkles and the like, can be used for improving the skin barrier function, relieving inflammatory response and delaying skin aging, and can be used for preventing and treating skin aging. The method can be widely applied to preparation of skin care products, cosmetics, hair supplies and daily chemical external preparations, and has good application prospects and market value. Meanwhile, the preparation method provided by the invention is mild in condition, high in reaction efficiency, environment-friendly and suitable for industrial production, and a raw material basis is provided for preparing more products for improving the skin barrier function, relieving inflammatory response and delaying skin aging.

The invention relates to a fluorescent probe compound for tumor targeted imaging and a synthesis method and application thereof.According to the compound, an anti-tumor drug with a pyrrolo [2, 3-d] pyrimidine core structure serves as a targeted ligand for the first time and is connected with a fluorescent dye after being modified by a connecting molecule, the structure of the compound is shown in the formula I, and the structural formula I is shown in the description. The method has high affinity and selectivity for tumor cells expressing target receptors in a targeted manner, can be used for in-vivo and in-vitro tracing of the tumor cells expressing the target receptors and positioning, qualitative and quantitative analysis of related experiments such as receptor affinity and action mechanisms, and has high specificity, sensitivity and intuition. The fluorescent probe disclosed by the invention can be quickly cleared in normal tissues and detained in tumor parts for a long time, so that the effect of in-vivo diagnosis can be achieved, and the fluorescent probe has a certain clinical application prospect and is applied to clinical intraoperative navigation.

The invention discloses an immobilized enzyme reaction application, and particularly relates to a method for catalytically synthesizing vitamin A palmitate in a solvent-free system by utilizing immobilized lipase. Through molecular modification, the specific enzyme activity of the lipase double mutant AF-R296P / L330S is maximally improved by 9.47 times, and under the condition that no other solvent is added, the reaction can still be carried out by taking molten methyl palmitate as a catalytic medium. The immobilized lipase is prepared by using the lipase mutant with improved activity and taking macroporous adsorption resin as a carrier, can efficiently catalyze vitamin A acetate and methyl palmitate to generate the vitamin A palmitate, and has a good application prospect in the production of the vitamin A palmitate.