28 results about "Enzyme structure" patented technology

(1) An artificial nucleic acid molecule comprising (a) at least one of a 3'-untranslated region (3'UTR) element and / or (b) a 5'-untranslated region (5'UTR) element, and (2) a poly(A) tail is described. The artificial nucleic acid molecule may further comprise a polypeptide comprising a reverse transcriptase (RT) domain and optionally an endonuclease domain for genome modification. A system comprising the artificial nucleic acid molecule and a method of using the system are also described.

The application discloses a glutamate decarboxylase multi-site combined mutant and application thereof. The application finds key sites by analyzing the enzyme structure and amino acid sequence of glutamate decarboxylase from Bifidobacterium breve and human GAD65, finally the key amino acid residue sites in the catalytic region and the vicinity of the active center are reformed (C168M, V167F, Y165S, F304Y, Y308F), so that the catalytic activity and specificity of the glutamate decarboxylase are regulated, the reaction type is changed from decarboxylation to incomplete oxidation, and the catalysis of glutamic acid forms the oxidation byproduct succinamic acid. In the process of GAD producing succinamic acid, the side reaction is enhanced, and the main reaction is weakened, so that the produced GABA is further oxidized into succinamic acid. The biological catalysis method can effectively improve the screening probability of the mutant amino acid sites, improve the experimental efficiency and feasibility, and screen the mutant enzyme with enzyme activity obviously superior to the parent enzyme.

The present disclosure relates to biocatalytic ligation methods for generating oligonucleotides and fusion polypeptides for use in the methods. In particular, the disclosure relates to biocatalytic ligation methods that incorporate ATP regeneration and fusion polypeptides that include a polyphosphate kinase domain and an ATP-dependent nucleic acid ligase domain.

The invention belongs to the technical field of antibodies, and discloses a nano antibody for hot start of Taq DNA polymerase and a construction method of the nano antibody. The nano-antibody is a monovalent nano-antibody for sealing a 3 '-5' excision enzyme structural domain of Taq DNA polymerase, a monovalent nano-antibody for sealing a 5 '-3' polymerization active structural domain of Taq DNA polymerase or a bivalent nano-antibody obtained by connecting the monovalent nano-antibody and the monovalent nano-antibody through flexible peptide; the amino acid sequence of the monovalent nano antibody for sealing the 3 '-5' excision enzyme structural domain of the Taq DNA polymerase is shown as SEQ ID NO: 1, the amino acid sequence of the monovalent nano antibody for sealing the 5 '-3' polymerization active structural domain of the Taq DNA polymerase is shown as SEQ ID NO: 2, and the amino acid sequence of the flexible peptide is shown as SEQ ID NO: 3.

Described herein are crystalline forms of small molecule inhibitor compounds (1) of Prolyl Hydroxylase Domain-containing Protein (PHD) and pharmaceutical compositions thereof, and methods of their use in the treatment of diseases or conditions that would benefit from treatment with inhibitors of Prolyl Hydroxylase Domain-containing Protein (PHD). (1)

The invention relates to the technical field of bioengineering, and discloses a glucose dehydrogenase mutant G321R and application thereof. The method comprises the following steps: firstly, carrying out molecular docking on glucose dehydrogenase and a glucose substrate to obtain a series of amino acid sites possibly influencing the enzyme structure and the combination of the enzyme and the substrate, then carrying out virtual saturated mutation on amino acids of the sites, and screening out mutation possibly enhancing the enzyme stability and activity through molecular dynamics simulation. A plurality of amino acid sites and mutation schemes capable of improving the stability and activity of the glucose dehydrogenase are obtained through screening after large-scale expression of the glucose dehydrogenase, the stability and activity of mutants obtained through single-point mutation and multi-point combined mutation of the sites are improved to different degrees, and the mutants can be used for preparing the glucose dehydrogenase mutant. The compound has high activity, can improve detection sensitivity and accuracy when used for detecting blood glucose, has high stability, is suitable for preparing kits and the like, and has wide application prospects.

Described are compositions comprising (1) at least one of: (a) a 3 '-untranslated region (3' UTR) element and / or (b) a 5 '-untranslated region (5' UTR) element; and (2) an artificial nucleic acid molecule with a poly (A) tail. The artificial nucleic acid molecule may further comprise a polypeptide comprising a reverse transcriptase (RT) domain and optionally an endonuclease domain for genome modification. Systems comprising the artificial nucleic acid molecules and methods of using the systems are also described.

Provided are a highly efficient base editing system, a preparation method therefor and the use thereof. Specifically, provided is a fusion protein, comprising a nuclease domain and an editing enzyme domain, wherein the editing enzyme domain is inserted into an insertion site in the nuclease domain. The nuclease domain in the fusion protein has the function of a nuclease, the editing enzyme domain in the fusion protein has the function of an editing enzyme, and the nuclease domain is derived from a Cas protein or a Cas protein mutant. On the basis of an activation-induced cytosine deaminase (AID), a new AID mutant having a shorter sequence and higher base editing efficiency is obtained by means of screening, and then the highly active AID mutant is randomly inserted into nSaCas9 (D10A) KKH, such that a new cytosine base editor EB-TAM having high base editing efficiency and an expanded and narrowed editing scope is successfully obtained by means of screening.

The invention relates to the technical field of bioengineering, and discloses a glucose dehydrogenase mutant S334M and application thereof. The method comprises the following steps: firstly, carrying out molecular docking on glucose dehydrogenase and a glucose substrate to obtain a series of amino acid sites possibly influencing the enzyme structure and the combination of the enzyme and the substrate, then carrying out virtual saturated mutation on amino acids of the sites, and screening out mutation possibly enhancing the enzyme stability and activity through molecular dynamics simulation. A plurality of amino acid sites and mutation schemes capable of improving the stability and activity of the glucose dehydrogenase are obtained through screening after large-scale expression of the glucose dehydrogenase, the stability and activity of mutants obtained through single-point mutation and multi-point combined mutation of the sites are improved to different degrees, and the mutants can be used for preparing the glucose dehydrogenase mutant. The compound has high activity, can improve detection sensitivity and accuracy when used for detecting blood glucose, has high stability, is suitable for preparing kits and the like, and has wide application prospects.

The invention relates to the technical field of bioengineering, and discloses a glucose dehydrogenase mutant S334K and application thereof. The method comprises the following steps: firstly, carrying out molecular docking on glucose dehydrogenase and a glucose substrate to obtain a series of amino acid sites possibly influencing the enzyme structure and the combination of the enzyme and the substrate, then carrying out virtual saturated mutation on amino acids of the sites, and screening out mutation possibly enhancing the enzyme stability and activity through molecular dynamics simulation. A plurality of amino acid sites and mutation schemes capable of improving the stability and activity of the glucose dehydrogenase are obtained through screening after large-scale expression of the glucose dehydrogenase, the stability and activity of mutants obtained through single-point mutation and multi-point combined mutation of the sites are improved to different degrees, and the mutants can be used for preparing the glucose dehydrogenase mutant. The compound has high activity, can improve detection sensitivity and accuracy when used for detecting blood glucose, has high stability, is suitable for preparing kits and the like, and has wide application prospects.

The application discloses a kind of nano bubble regulated hydrogen bond organic framework immobilized enzyme, and hydrogen bond organic framework immobilized enzyme is prepared by in-situ self-assembly and embedding method in water environment containing nano bubble from four (4-amidinophenyl) methane and four (4-carboxyphenyl) methane.The hydrogen bond organic framework is used to realize the immobilization of enzyme by in-situ embedding method, and a multi-level pore immobilized enzyme system based on nano bubble is constructed.Nano bubble can expand the pore size and pore volume of hydrogen bond organic framework as pore-forming template, while maintaining the stability of enzyme structure, significantly improving the mass transfer rate of substrate molecules in the framework, thereby enhancing the catalytic activity and substrate affinity of immobilized enzyme.The method effectively improves the enzyme catalytic reaction efficiency while maintaining the system immobilization efficiency, pH stability, temperature stability and reusability.The process is simple and suitable for the construction of multi-level pore immobilized system for various enzymes.

Provided herein are fusion proteins comprising transposase domains and DNA targeting domains. In particular, the DNA targeting domains may be targeted to the lipoprotein A (LPA) gene. Also provided are methods of making the transposase domains and fusion proteins, cells that are modified using the fusion proteins provided herein and methods of treatment using such cells.

The invention discloses a composite protective agent for liquid laccase and application of the composite protective agent, and belongs to the technical field of bioengineering. The protective agent takes a citric acid-disodium hydrogen phosphate buffer solution with the pH value of 4.5-6.5 as a solvent, and comprises propylene glycol, glycerol, sorbitol, cane sugar, maltodextrin, glycine, trehalose, potassium sorbate, diacetic acid and a mediator. The components form a stable microenvironment around enzyme molecules through a synergistic effect, the polyhydric alcohols and the saccharides stabilize an enzyme structure through hydrogen bonds and glassy state protection, the glycine plays a buffering role, and the mediator has dual functions of oxidation resistance and mediated catalytic reaction. Experiments prove that the protective agent can remarkably improve the thermal stability and storage stability of the liquid laccase, the enzyme activity retention rate reaches 86.7% after the liquid laccase is treated at 50 DEG C for 24 h, and the enzyme activity retention rate is still kept at 85.3% after the liquid laccase is stored at 37 DEG C for 180 days. The laccase is especially suitable for degrading aflatoxin B1, has a degradation effect obviously better than that of unprotected laccase, and can be widely applied to the fields of food industry, feed industry and the like.

Conjugation reagents for making, and targeted degrader conjugates made therefrom, comprising a Janus family kinase binder and a CD127 binding agent, are described. In particular, the conjugation reagents are compounds of Formula (I) where TM is a JANUS family kinase domain-targeting binder; L1 is a first linker; L2 is a second linker; and E3L is a E3 ligase ligand; wherein, TM, L2 and E3L are covalently linked to L1, and L2 comprises a reactive moiety for reacting with a binding molecule. (I)

This disclosure generally relates to transposase domains, in particular, transposase domains comprising amino terminal deletions, as well as transposase domains forming obligate heterodimers and transposase domains comprising DNA targeting domains.

This invention proposes a method for screening and optimizing enzyme catalytic efficiency and stability based on protein language models and domain knowledge. The method includes: defining a set of allowable mutation intervals through multidimensional analysis; constructing a dual-source mutant library using a dual-source complementarity strategy based on protein language models and co-evolutionary models; using the dual-source mutant library as the initial population, employing a multi-model fusion catalytic efficiency prediction strategy as the population's overall fitness calculation method, and a dual-index verification strategy based on the SPIRED-Stab model for structural stability as the population screening method; iteratively obtaining the optimal mutant enzyme sequence through a semantically guided genetic algorithm; and verifying the structure and dynamic stability of the optimal mutant enzyme sequence based on three-dimensional structure prediction and molecular dynamics simulation to obtain an experimentally feasible final mutant enzyme sequence. This invention achieves improved optimization efficiency while ensuring the structural feasibility of key enzymes.

The invention relates to a structural analysis-based rational transformation method for improving enzymatic properties of beta-mannase under an alkaline condition, and belongs to the technical field of enzyme engineering and industrial biology. The method comprises the following steps: on the basis of a three-dimensional structure model of beta-mannase, identifying aromatic amino acid residues which are positioned in a catalytic center or near a substrate binding channel and play an important role in enzyme conformation stability and catalytic function; on the basis, establishing a negative limitation principle of rational design of catalytic activity, namely avoiding amino acid replacement of the aromatic amino acid residues in an enzyme engineering design process; on the premise of following the negative limitation principle, a substrate binding structure domain of the beta-mannase is constructed in series, so that the enzyme structure stability and the substrate binding capacity are synergistically improved, and the beta-mannase with improved stability and catalytic performance under the alkaline condition is obtained. According to the method, blind mutation is avoided, a rational design method for improving the enzymatic performance of the beta-mannase under the alkaline condition is provided, the rational design method has a clear structural basis and an engineering modification path, and the method has a good industrial application prospect.

The invention provides an efficient base editing system and a preparation method and application thereof. Specifically, the invention provides a fusion protein which comprises a nuclease structural domain and an editing enzyme structural domain, wherein the editing enzyme structural domain is inserted into an insertion site in the nuclease structural domain, and the nuclease structural domain in the fusion protein has a nuclease function; the editing enzyme structural domain in the fusion protein has an editing enzyme function; the nuclease structural domain is derived from a Cas protein or a Cas protein mutant. According to the present invention, based on activation induction cytosine deaminase AID, a novel AID mutant with characteristics of short sequence and high base editing efficiency is screened, and then the high-activity AID mutant is randomly inserted into nSaCas9 (D10A) KKH so as to successfully screen the novel cytosine base editor EB-TAM with characteristics of high base editing efficiency, wide editing range and narrow editing range.

The present application relates to the technical field of bioengineering, and discloses a glucose dehydrogenase mutant with enhanced stability and activity; firstly, molecular docking is carried out on glucose dehydrogenase and glucose substrate to obtain a series of amino acid sites which may affect enzyme structure and enzyme-substrate binding; then, virtual saturation mutation is carried out on the amino acids of the sites; through molecular dynamics simulation, mutations which may enhance enzyme stability and activity are screened; after starting wet experiment and screening through large-scale expression, multiple amino acid sites and mutation schemes which improve the stability and activity of glucose dehydrogenase are obtained; the mutants obtained through single-point mutation and multi-point combined mutation of the sites have different degrees of improvement in stability and activity; the high activity is used for detecting blood glucose, and can improve detection sensitivity and accuracy; the high stability is suitable for preparing reagent kits, and has a wide application prospect.

The invention provides a construction method and application of a P450 fusion protein. The construction method comprises the following steps: step (1), fusing abamectin oxidation type P450 Ema1 with a Rhodococcus P450RhF-RPs reductase structural domain by adopting a connecting peptide to obtain Ema1-Rhf; and (2) carrying out point mutation on the Ema1-Rhf, and constructing the P450 fusion protein. According to the invention, redox partners derived from Rhodococcus are connected with a structural domain of P450 Ema1 through a designed connecting peptide; the cytochrome P450 monooxygenase mutant further improves the enzyme activity in the process of catalyzing abamectin to be converted into methylamino abamectin, the maximum conversion rate can be improved by 2.2 times through determination, and the mutant achieves 90% conversion rate within 39.8 hours, which is far better than the expression of an unfused system. And the method has certain significance on industrial production of the emamectin benzoate.

The present invention belongs to the technical field of the preparation of fused enzymes. Disclosed are a eurythermal, broad-pH and salt-tolerant dual-enzyme-coupled recombinant fused β-mannanase and the use thereof. For the first time, a computer-aided molecular design is used to couple a cold-active enzyme from Bacillus clausii to a protein sequence-modified thermophilic enzyme from Thermotoga maritima via linker L, and to add a C-terminal polypeptide sequence for improving the structural stability of the enzyme at the C-terminus, and a fused enzyme is obtained by means of the induced expression of constructed recombinant Escherichia coli. The obtained fused enzyme can tolerate a pH value of 4-11, a temperature of 20-100°C, and salinity of up to 20%, has the characteristic of initiating an enzymatic reaction at both low and high temperatures, respectively, and can be widely used in pulp bleaching, detergents, foods, feeds, coffee processing, petroleum exploitation, drug development, biofuel production, and the enzymatic production of mannooligosaccharides. The enzyme can simplify the problem of the simultaneous requirement for different types of β-mannanases in the production process, can be used for multiple purposes, and has broad application prospects.

A computer-implemented method for predicting quantum tunnelling enhancements in enzyme reaction rates. An enzyme structure is retrieved from a public database such as the Protein Data Bank (PDB). A qu

The invention discloses a method convenient for plant pilot editing, which comprises the following steps: respectively carrying out cis-connection on a Cmylc promoter and two core editing elements through a restriction enzyme site directional cloning technology; a Cmylc promoter-nCas9-RT expression unit is constructed on the upstream of a gene (the sequence comprises a nuclear localization signal peptide coding region, an nCas9 nicking enzyme structural domain coding region and a reverse transcriptase functional domain coding region) for coding an nCas9-RT fusion protein, and it is ensured that transcription initiation of the fusion protein is regulated and controlled by a Cmylc promoter; the editing efficiency is enhanced from two dimensions by utilizing the unique transcriptional regulation ability of the Cmylc promoter in tobacco cells, on one hand, by improving the transcriptional level of nCas9-RT fusion protein and increasing the accumulation amount of functional fusion protein in cell kernels, a sufficient substrate is provided for assembly of an editing compound (nCas9-RT / pegRNA), and on the other hand, the editing efficiency is improved from two dimensions; finally, through cooperative high expression of'protein-nucleic acid 'double elements, the core limitation of'insufficient element expression quantity' in a traditional editing system is broken through, and the order of magnitude improvement of the tobacco leading editing efficiency is achieved.

The invention relates to the technical field of bioengineering, and discloses a glucose dehydrogenase mutant S413V and application thereof. The method comprises the following steps: firstly, carrying out molecular docking on glucose dehydrogenase and a glucose substrate to obtain a series of amino acid sites possibly influencing the enzyme structure and the combination of the enzyme and the substrate, then carrying out virtual saturated mutation on amino acids of the sites, and screening out mutation possibly enhancing the enzyme stability and activity through molecular dynamics simulation. A plurality of amino acid sites and mutation schemes capable of improving the stability and activity of the glucose dehydrogenase are obtained through screening after large-scale expression of the glucose dehydrogenase, the stability and activity of mutants obtained through single-point mutation and multi-point combined mutation of the sites are improved to different degrees, and the mutants can be used for preparing the glucose dehydrogenase mutant. The compound has high activity, can improve detection sensitivity and accuracy when used for detecting blood glucose, has high stability, is suitable for preparing kits and the like, and has wide application prospects.

The application discloses a cobalt-based porous chiral metal-organic framework enzyme-mimic material and a preparation method and application thereof, relates to the technical field of chiral science, and the material is formed by one-pot coordination assembly with chiral H4L as a bridging ligand, 4, 4'-dipyridyl as an auxiliary ligand and cobalt as a metal salt; the application also provides a cobalt-based porous chiral metal-organic framework enzyme-mimic material prepared by the method and application thereof. The cobalt-based porous chiral metal-organic framework with unique cubic cavities, narrow open channels and chiral recognition sites and the enzyme-mimic structure characteristics can be assembled by coordination of the chiral ligand H4L, 4, 4'-dipyridyl and cobalt ions; the preparation method is simple, the conditions are mild and controllable, the yield is high and the method is easy to repeat. The material can separate enantiomers of chiral secondary alcohols with excellent enantioselectivity, size selectivity, substrate universality and recycling.

The invention discloses a complex microbial inoculant as well as a preparation method and application thereof. The complex microbial inoculant contains stenotrophomonas maltophilia and bacillus stratophilus. Wherein the enzyme activity of the stratospheric bacillus is relatively high, and the stratospheric bacillus can efficiently catalyze the hydrolytic decomposition of organic matters; the stenotrophomonas maltophilia has strong capability of producing active oxygen, and can effectively destroy the chemical structure of pollutants so as to improve the degradation efficiency of the pollutants. Active oxygen generated by stenotrophomonas maltophilia can serve as a signal molecule to activate or regulate the enzyme activity through a reversible modification enzyme structure, meanwhile, the high enzyme activity of the bacillus stratophilia can also reversely regulate the active oxygen level, the active oxygen and the bacillus stratophilia can form a core regulation relation of dynamic balance and mutual promotion, functional characteristics are precisely complementary, and the activity of the active oxygen is improved. And thus, synchronous and efficient degradation of various pollutants is realized. Moreover, strains used by the complex microbial inoculant are commercially available standard strains, the sources are clear, the preparation method is simple and controllable, and the complex microbial inoculant has excellent industrial application prospects.

The invention relates to the technical field of bioengineering, and discloses a glucose dehydrogenase mutant S503P and application thereof. The method comprises the following steps: firstly, carrying out molecular docking on glucose dehydrogenase and a glucose substrate to obtain a series of amino acid sites possibly influencing the enzyme structure and the combination of the enzyme and the substrate, then carrying out virtual saturated mutation on amino acids of the sites, and screening out mutation possibly enhancing the enzyme stability and activity through molecular dynamics simulation. A plurality of amino acid sites and mutation schemes capable of improving the stability and activity of the glucose dehydrogenase are obtained through screening after large-scale expression of the glucose dehydrogenase, the stability and activity of mutants obtained through single-point mutation and multi-point combined mutation of the sites are improved to different degrees, and the mutants can be used for preparing the glucose dehydrogenase mutant. The compound has high activity, can improve detection sensitivity and accuracy when used for detecting blood glucose, has high stability, is suitable for preparing kits and the like, and has wide application prospects.

The invention relates to the technical field of food processing, and discloses a freeze-dried noodle processing system adopting an enzyme deactivation structure, the freeze-dried noodle processing system comprises a placing member, the placing member comprises a fixing frame slidably connected with the top end of a supporting seat, outer trays are vertically stacked on the inner wall of the fixing frame, the inner walls of the outer trays are symmetrically provided with limiting grooves, and the inner walls of the limiting grooves are slidably connected with limiting blocks; the other sides of the symmetrically-arranged limiting blocks are fixedly connected with inner supporting discs. The inner supporting disc is arranged, the convex strips on the inner wall of the hole and the supporting strips at the intervals form a multi-point uniform supporting structure, and compared with a traditional single-point hanging mode, the design can disperse the gravity of noodles to a plurality of contact points, breakage caused by too large local tension is avoided, and the service life of the noodles is prolonged. And meanwhile, the convex strips are made of flexible materials and can be elastically attached to the surfaces of the noodles, so that the microstructures of the noodles are not damaged, and stable supporting force can be provided.