270 results about "Amino acid mutation" patented technology

The invention discloses a Taq DNA polymerase mutant as well as a preparation method and application thereof, and relates to the technical field of biology. The Taq DNA polymerase mutant is mutation on the basis of a wild type Taq DNA polymerase amino acid sequence as shown in SEQ ID NO.1, and comprises at least one of amino acid mutation L345E, L352D and S438M. The Taq DNA polymerase mutant has excellent tolerance to urea, and solves the dual technical problems that complex biological samples containing endogenous inhibitors such as urea can be efficiently amplified, and the Taq DNA polymerase mutant is compatible with difficult PCR reactions which take urea as an additive to optimize a high GC template and the like. Therefore, the mutant is a molecular biology tool enzyme which is wider in applicability and more stable and reliable in performance.

The invention discloses a quinoa CqARF2 gene mutant as well as an identification method and application thereof. Chenopodium quinoa seeds are mutated on a large scale by utilizing a chemical mutagenic agent EMS, a CqARF2 gene function deletion mutant arf2 is adopted as a quinoa mutant, G at the 1050th site of the CqARF2 gene of the mutant is mutated into A, and amino acid at the 350th site is mutated into a termination codon, so that the translation process of CqARF2 protein is terminated in advance. The chenopodium quinoa CqARF2 gene function deletion mutant arf2 shows remarkable agronomic character improvement, seeds are large and full, compared with a wild type, the hundred-grain weight of the mutant seeds is increased by about 40%, the seed diameter is increased by about 10%, the seed thickness is increased by about 30%, and a certain foundation is laid for breeding new high-yield and high-quality chenopodium quinoa varieties.

The invention relates to the field of tumor cell therapy, in particular to application of T lymphocyte chimeric with a B7-H3 receptor to treatment of head and neck tumors, and provides an anti-B7-H3 scFv, the scFv comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region comprises an HCDR region and an HFR region, and the light chain variable region comprises an LCDR region and an LFR region; compared with a wild type scFv sequence, the scFv sequence has the advantage that a plurality of amino acids with positive charges in the HFR region and / or the LFR region are mutated into amino acids without charges. According to the invention, positive charge plaques on the CAR surface of an scFv sequence of a B7-H3 human-derived monoclonal antibody MGA271 are changed in a charged amino acid mutation manner, so that a B7-H3. CAR-T cell is optimized, and it is proved that the B7-H3. CAR-T cell optimized by PCP can effectively kill B7-H3 positive tumor cells in vivo and in vitro; and a new method and thought are provided for clinical targeted treatment of B7-H3 positive solid tumors.

The present invention provides an α-amino acid ester acyltransferase mutant and its application. The α-amino acid ester acyltransferase mutant includes: (a) a protein having the amino acid sequence shown in SEQ ID NO: 1; (b) a protein having an amino acid mutation at at least one of the following sites in the amino acid sequence in (a): P158, K80, T208, A302, A175, N85, N605, K346, S225, I34, or S348, and having the function of an α-amino acid ester acyltransferase; or (c) a protein having more than 80% homology with the amino acid sequence defined in (a) or (b) and having the function of an α-amino acid ester acyltransferase. The α-amino acid ester acyltransferase mutant of the present invention has the advantages of a broad substrate spectrum and high catalytic activity, can be well used in the synthesis of oligopeptides, has low cost and high yield, and realizes true green chemistry.

The invention belongs to the technical field of bioengineering, and particularly relates to a zearalenone toxin degrading enzyme variant and application thereof. The amino acid sequence of the zearalenone toxin degrading enzyme is as shown in SEQ ID NO.2, or the zearalenone toxin degrading enzyme with the amino acid sequence as shown in SEQ ID NO.2 is obtained through amino acid mutation. The zearalenone toxin degrading enzyme mutant is obtained through mutation screening, and compared with a wild type, the zearalenone toxin degrading enzyme mutant has the advantages that the zearalenone degrading capability is obviously improved; besides, the zearalenone toxin degrading enzyme is expressed by using alfalfa, the zearalenone toxin degrading enzyme with biological activity is easy to obtain, and the zearalenone toxin degrading enzyme has certain application potential in prevention and treatment of animal poisoning caused by zearalenone toxin pollution in agriculture and animal husbandry production.

Disclosed herein are predictive models for T-cell epitope prediction, B-cell epitope prediction, and protein design wherein a method is implemented for generating a protein variant amino acid sequence of a target protein having one or more modified properties, the method comprising: (a) iteratively sampling an input amino acid sequence of the target protein, and (b) sampling the individual protein score of at least one weighted relative contribution of the single residue mutant input amino acid sequence to the at least one target property across a plurality of other single residue mutant input amino acid sequences to generate a combined protein score, wherein the combined protein score corresponds to the protein variant comprising one or more amino acid mutations of the single residue mutant input amino acid sequences.

The present invention relates to the technical field of biomedicine. Specifically provided are a respiratory syncytial virus (RSV) Pre-F recombinant protein nanoparticle vaccine and a preparation method therefor. By performing amino acid mutation on an RSV Pre-F protein, and performing fusion expression on the mutated Pre-F protein and ferritin particles in eukaryotic cells, PreF-Ferritin fusion protein nanoparticles, in which eight Pre-F protein trimers are densely displayed on the surface of each ferritin particle, are obtained. By stabilizing and exposing antigen epitopes required to be displayed and disrupting or masking unrequired antigen epitopes, the PreF-Ferritin fusion protein nanoparticles effectively improve the immunogenicity and the production stability of the antigen. Experiments show that: when the PreF-Ferritin fusion protein is injected into mice, high-titer protective sera can be acquired, and the mice sera can yield relatively high neutralizing titers against RSV.

A varicella-zoster virus (VZV) nanoparticle protein, and a preparation method therefor and a use thereof. The VZV protein comprises a partial or full sequence of an amino acid sequence of an extracellular region of a VZV gE glycoprotein, with W at position 200 mutation to C and L at position 245 mutation to C in the amino acid sequence of the extracellular region of the VZV gE glycoprotein. By means of the rational optimization design of the amino acid sequence of the VZV gE protein by means of protein genetic engineering, the VZV gE recombinant protein modified with amino acid mutations has increased stability and immunogenicity compared with the VZV gE protein. Moreover, by further designing the protein structure, the VZV gE protein is repeatedly displayed on the surface of ferritin nanoparticles with the desired epitopes exposed, thereby further enhancing immunogenicity.

The invention discloses an anti-inhibition Taq DNA polymerase and application thereof, and relates to the technical field of biology. According to the anti-inhibition Taq DNA polymerase, amino acid mutations of K206R, F667Y and the like are carried out on an amino acid sequence of a wild type Taq DNA polymerase with an amino acid sequence as shown in SEQ ID No.1, so that a certain degree of anti-inhibition property is obtained. In addition, the invention also discloses application of the anti-inhibition Taq DNA polymerase in preparation of reagents or kits, and the like. Compared with a wild type Taq DNA polymerase, the anti-inhibition Taq DNA polymerase can maintain the PCR amplification capability in the presence of an inhibitor such as polysaccharide.

The invention provides a glutaminase mutant and application of the glutaminase mutant. Wherein the glutaminase mutant comprises (a) a protein which is subjected to amino acid mutation and has a glutaminase activity function on at least one of the following sites of S467, Y334, K400, N521, R337, E402 or D447 of an amino acid sequence as shown in SEQ ID NO: 2, and (b) a protein which is subjected to amino acid mutation on at least one of the following sites of S334, Y334, K400, N521, R337, E402 or D447 of the amino acid sequence as shown in SEQ ID NO: 2; and (b) a protein which has 80% or more homology with the amino acid sequence limited by (a) and has a glutaminase function. The glutaminase mutant with the amino acid sequence disclosed by the invention has relatively high specific enzyme activity during enzyme catalytic reaction, and is low in cost and strong in market competitiveness when being used for commercial food processing.

A cage-like nanocarrier for targeted delivery of siRNA, and a preparation method therefor and the use thereof. The preparation method comprises: (A) mutating a negatively charged or uncharged amino acid on the inner surface of a ferritin into a positively charged amino acid; and any one or more of the following steps: (B) coupling the N-terminus of the ferritin to a functional peptide having nucleic acid affinity; (C) coupling the N-terminus of the ferritin to a functional peptide promoting lysosomal escape; (D) truncating the E-helix at the C-terminus of the ferritin; (E) coupling the C-terminus of the ferritin to a functional peptide having nucleic acid affinity; and (F) coupling the C-terminus of the ferritin to a functional peptide promoting lysosomal escape. A new nucleic-acid-loaded protein nanocage carrier is constructed by means of modifying a negatively charged inner cavity of ferritin to make same positively charged. By means of electrostatic adsorption, a negatively charged siRNA can be efficiently loaded into a ferritin nanocage, thereby significantly improving the in-vivo and in-vitro delivery stability of siRNA, lysosomal escape functions, and efficacy of targeted therapy.

The invention belongs to the technical field of target sample characterization, and provides application of enterococcus pore-forming toxin mutant protein in nanopore detection. The enterococcus pore-forming toxin mutant protein is EPX1 nanopore mutant protein and is obtained by mutating wild type EPX1 nanopore protein, the amino acid sequence of the wild type EPX1 nanopore protein is as shown in SEQ ID NO: 1, and the mutation means that one or more amino acids of the wild type EPX1 nanopore protein are mutated into common amino acids except original amino acids. The EPX1 protein (wild type) is a type of pore-forming toxin protein and widely exists in various living bodies in nature, and a mutant of the EPX1 protein shows excellent pore-forming capacity after being modified (after amino acid mutation). The structural stability, the molecular recognition efficiency and the detection performance of the EPX1 nanopore mutant protein are remarkably improved.

This invention belongs to the field of enzyme engineering technology and discloses a BhrPETase mutant and its applications. The invention provides a BhrPETase mutant with the amino acid sequence shown in SEQ ID No. 2, designated as mutant F208I. Compared to the wild-type mutant BhrPETase, this mutant exhibits a 1.39-fold increase in PET degradation activity. Based on mutant F208I, this invention further rationally designs amino acid mutations, providing 21 BhrPETase mutants. The BhrPETase mutants provided by this invention, while maintaining thermal stability essentially equivalent to wild-type BhrPETase, increase PET degradation activity by 1.39 to 4.24 times. The BhrPETase mutants provided by this invention can be applied in fields such as PET degradation, PET degradation product recovery, or the preparation of PET degradation agents.

The invention provides a method for preparing (R)-4-chloro-3-hydroxybutyric acid ethyl ester ((R)-CHBE) through enzyme catalysis, which comprises the following steps: isopropanol is used as a hydrogen donor, a ketoreductase mutant is used for carrying out catalytic reduction reaction of COBE to prepare (R)-CHBE, and the ketoreductase mutant has amino acid mutation at the 83rd site on the basis of an amino acid sequence shown as SEQ ID NO.1. The invention also provides a preparation method of (R)-4-chloro-3-hydroxybutyric acid ethyl ester ((R)-CHBE). The thermal stability, namely heat resistance, of the ketoreductase mutant is improved, and by combining with the use of a rectifying tower, the method disclosed by the invention improves the utilization rate of isopropanol and can discharge a byproduct acetone in time, so that the substrate conversion rate and the product concentration are improved. The invention also provides an enzyme catalysis device used in the method.

The invention relates to the technical field of biological detection, and particularly discloses an ELISA kit for detecting a neutralizing antibody of a severe fever with thrombocytopenia syndrome virus and an application method of the ELISA kit. The ELISA kit comprises an ELISA plate on which SFTSV receptor LRP1 protein is fixed, and an enzyme-labeled SFTSV Gn-8M, wherein the SFTSV receptor LRP1 protein is fixed on the ELISA plate; the SFTSV Gn-8M is an SFTSV membrane protein Gn mutant designed by the invention, and can be specifically combined with an SFTSV receptor LRP1 protein on an elisa plate, and compared with a standard virus strain Gn protein, the self-designed SFTSV Gn-8M has amino acid mutation of eight sites, has stronger combining capacity with a host cell receptor, and can be used for detecting the SFTSV. The kit disclosed by the invention has the characteristics of high safety, high efficiency, high flux, high specificity, high sensitivity, high stability and the like, and is suitable for large-scale rapid screening of the neutralizing antibody of the severe fever with thrombocytopenia syndrome virus.

The invention discloses a milk-derived broad-spectrum antibacterial peptide W-2 and application thereof, and relates to the technical field of biology. The amino acid sequence of the antibacterial peptide is as shown in SEQ ID NO. 4. Symmetrical structure design and amino acid mutation are carried out on the basis of the template peptide to obtain the novel antibacterial peptide, and tests show that the novel antibacterial peptide has broad-spectrum antibacterial activity which is obviously higher than that of the template peptide, and basically has no cytotoxicity. The antibacterial peptide prepared by the invention still keeps good antibacterial activity in physiological salt concentration and protease environment, has certain serum stability, and has higher application value; in addition, the antibacterial peptide has a good inhibition effect on the formation of a bacterial biofilm, and has a synergistic effect with traditional antibiotics.

The invention belongs to the technical field of biology, and particularly relates to an attenuated porcine epidemic diarrhea virus recombinant strain, a construction method and application. According to the invention, amino acid N at the 381th site of PEDV S protein is mutated into K, and a recombinant strain rPEDV-SN381K is obtained through rescue; compared with a parent wild virus rPEDV-Swt, the recombinant virus strain rPEDV-SN381K has the advantages that the virulence of the recombinant virus strain rPEDV-SN381K is obviously weakened, the infection of the rPEDV-Swt can be effectively resisted, and the recombinant virus strain rPEDV-SN381K can be used for preparing a PEDV vaccine.

The invention discloses an Apopsin3 mutant and a method for regulating an environment in a plant cell to be acidic, and belongs to the technical field of pH regulation in the plant cell. The regulation and control method comprises the following steps: mutating 83rd and 120th amino acids of the light-operated hydrogen ion pump protein Apopsin3, converting into hydrogen ion channels, expressing the hydrogen ion channels on a cell membrane in a targeting manner, and stimulating with green light of 40-500 [mu] W / mm < 2 > and 520-560 nm so as to regulate the pH in the cell to be acidic. By adopting the cell specific promoter and performing site-specific mutagenesis modification on the specific hydrogen ion pump, accurate pH regulation and control on specific cells or organelles can be realized. An Apopsin3 (FARH) gene is fused with a cell membrane targeting sequence, and under the stimulation of specific light, only the pH of cytoplasm is obviously changed, and the pH of other cell regions is not influenced.

The invention provides a hyoscyamine 6 beta hydroxylase mutant as well as a preparation method and application thereof. The hyoscyamine 6 beta hydroxylase mutant at least has the following two sites or amino acid mutation in functional equivalent sites compared with wild hyoscyamine 6 beta hydroxylase: the 14th site and the 97th site; the wild type hyoscyamine 6 beta hydroxylase has an amino acid sequence as shown in SEQ ID NO: 1. The obtained hyoscyamine 6 beta hydroxylase mutant has higher hydroxylation activity, and meanwhile, the epoxidation capacity disappears or is reduced. In industrial application, the catalytic efficiency can be remarkably improved, reagent waste is reduced, the production cost is saved, generation of redundant by-products in chemical reaction is reduced, and environmental pollution is reduced.

The invention discloses an XXM-ER mutant for regulating and controlling plant endoplasmic reticulum calcium ion homeostasis, nucleic acid and application, and belongs to the technical field of biological engineering. The XXM-ER mutant has two amino acid mutations of H134Q and D156H on a light-operated cation channel protein ChR2, and the amino acid sequence of the mutant is as shown in SEQ ID NO. 1. The XXM-ER mutant disclosed by the invention has better endoplasmic reticulum localization, can realize precise regulation and control of plant calcium ion flow through illumination regulation and control, and has great application value in the aspects of plant cultivation, plant calcium signal research and the like.

The invention relates to the field of biochemical engineering, in particular to an alcohol dehydrogenase mutant and application thereof. The mutant is obtained by carrying out mutation on one or more sites selected from 111Y, 258V, 283L, 274L, 107K and 108N aiming at alcohol dehydrogenase, and the mutation is that amino acid at the sites is mutated into one of W, Q, V, G and A. The enzyme activity of the alcohol dehydrogenase mutant is higher than that of a wild type; the enzyme specific activity (36.42) of the mutant A-111-107-274 taking FAL as a substrate is 2.4 times that of a wild type, and the enzyme specific activity (22.16) of the mutant A-111-107-274 taking HMF as a substrate is 9.2 times that of the wild type. Good industrial application performance is shown.

The invention relates to the field of biological medicine, and provides a coxsackie virus A16 type mutant and a virus-like particle thereof, compared with a wild type coxsackie virus A16 type, the mutant has at least one amino acid mutation site in the following regions: (1) a canyon region of a virus capsid; (2) a channel area at a secondary axis; or (3) a channel region at a quasi-tertiary axis. The mutant and the virus-like particle are obtained by optimizing and modifying structural proteins VP0, VP1 and VP3 which form a virus capsid by using computational biology and structural biology methods, co-expressing and co-assembling the structural proteins in a hansenula polymorpha expression system to form the virus-like particle, and then carrying out series of chromatographic purification. Compared with a non-mutated coxsackie virus A16 type virus-like particle mutation scheme, the coxsackie virus A16 type virus-like particle mutation scheme disclosed by the invention has the advantages that the antigen immunogenicity can be obviously improved, and the clinical application value is realized.

The invention provides a DNA polymerase mutant and application thereof. Compared with a wild type KOD DNA polymerase, the DNA polymerase mutant has amino acid mutation at the 408th site, the 409th site or a functional equivalent site; and an amino acid mutation of at least one site selected from the following 23 sites and functional equivalence sites: 141 site, 143 site, 147 site, 383 site, 384 site, 389 site, 485 site, 584 site, 589 site, 397 site, 424 site, 432 site, 445 site, 523 site, 553 site, 561 site, 564 site, 461 site, 481 site, 605 site, 663 site, 711 site, and 725 site; the amino acid sequence except the amino acid mutation site of the mutant has at least 90% identity with the corresponding amino acid sequence of the wild type KODDNA polymerase; the wild type KOD DNA polymerase has an amino acid sequence as shown in SEQ ID NO: 2.

The present invention provides a carbonyl reductase mutant, preparation method and use thereof, and a preparation method of ethyl (R)-6-hydroxy-8-chlorooctanoate. The carbonyl reductase mutant is a carbonyl reductase with amino acid mutation; the carbonyl reductase comprises an amino acid sequence as set forth in SEQ ID NO: 2; the amino acid mutation includes E101V, F214R or E101V / F214R. In the present invention, by introducing mutations on the basis of the original carbonyl reductase sequence, the enzyme activity is improved, the stereoselectivity is improved, and ethyl (R)-6-hydroxy-8-chlorooctanoate can be obtained with high yield and high purity under relatively mild conditions, which reduces the production cost and is suitable for industrial production.

The present invention discloses an L-arginine-glycine amidinotransferase and use thereof in the production of guanidinoacetic acid. In the present invention, through combined multi-site amino acid mutation, a technical effect of significantly improved enzyme activity of the mutant AkAGATT225Q / A258P / L278K than that of the wild-type strain is achieved, providing an application value for large-scale production of guanidinoacetic acid in industry. When the L-arginine-glycine amidinotransferase mutant constructed in the present invention is used in the production of guanidinoacetic acid, by optimizing the conversion conditions, the yield of guanidinoacetic acid is up to 21.4 g / L and the conversion rate is 90.4%, after 24 hrs of reaction in a 1 L reaction system. Compared with the production of guanidinoacetic acid with the raw enzyme, the yield is increased by 49.6%.

The invention relates to the technical field of enzyme engineering, in particular to a xylose reductase XR21 mutant and application thereof. Specifically, the invention provides a novel xylose reductase mutant aiming at the defect of poorer thermal stability of the existing wild type xylose reductase, and particularly provides a xylose reductase mutant which is obtained by performing single or multiple site amino acid mutation on the 19th site, the 48th site, the 79th site, the 208th site and the 300th site of a xylose reductase sequence shown as SEQ ID NO: 2, compared with wild type xylose reductase, the obtained xylose reductase mutant has the advantages that the activity of catalyzing an epimer substrate arabinose into arabitol can be obviously reduced, and meanwhile, the selectivity to xylose is improved, so that the operation process of downstream separation and purification is simplified, and the biological catalysis requirement of green industry is met.

The invention discloses a double-function (p) ppGpp synthetase or hydrolase mutant, a biological material and application of the double-function (p) ppGpp synthetase or hydrolase mutant to preparation of L-threonine, and belongs to the technical field of gene engineering. The technical problem to be solved by the invention is how to improve the yield of L-threonine. Relative to a wild type, the bifunctional (p) ppGpp synthetase or hydrolase mutant comprises one or more of the following mutations: K1) mutation of 174th amino acid of SEQ ID NO: 2 into cysteine, K2) mutation of 529th amino acid of SEQ ID NO: 2 into phenylalanine, and K3) insertion of histidine and aspartic acid between 84th amino acid and 85th amino acid of SEQ ID NO: 2. The bifunctional (p) ppGpp synthetase or hydrolase mutant provided by the invention can be used for improving the yield of L-threonine of Escherichia coli.

The application discloses a key type specificity changed alpha-glucosyltransferase mutant, and belongs to the technical field of enzyme engineering and starch modification. The method is characterized in that, by analyzing the combination of Lf2970GtfB 4,3-alpha-glucosyltransferase in the active center donor / acceptor sub-site and the substrate, part of key amino acids of Lf2970GtfB are mutated into different types of amino acids, the mutant is prepared by changing the enzyme and substrate interaction force, space steric hindrance and other factors affecting the orientation of the substrate in the active center, the transglycosylation is changed, and the bond type composition of the product is changed. The application changes the bond type specificity of Lf2970GtfB 4,3-alpha-glucosyltransferase by mutation, the obtained mutant has novel 4,3 / 6-alpha-glucosyltransferase activity, and is helpful to synthesizing unique structure and more valuable alpha-glucan by using starch substrates.

This invention belongs to the field of target characterization technology, and provides an application of PANX2 protein and its mutants as bioporins. The bioporins can detect one or more of the following: metal ions, inorganic salts, amino acids, nucleotides, small molecule drugs, small molecule diagnostic reagents, adenosine triphosphate (ATP), and monosaccharides. In this invention, one or more amino acids of the wild-type PANX2 protein are mutated to common amino acids other than the original amino acids. Compared to the wild-type PANX2 protein, the PANX2 protein mutants can be expressed normally, are stable, and show a significantly improved current signal during detection, with a longer amplitude and fewer spikes.

The application discloses a D type influenza virus isolate D / HY11 and application thereof in a mink infection model, and belongs to the field of biological medicines. The application takes the D type influenza virus isolate D / HY11 of an IDV northeast isolate as a research object, establishes a mink transmission model, evaluates the transmission dynamics characteristics of the strain among minks, identifies amino acid mutations possibly related to host adaptability through whole genome sequencing, analyzes the amino acid substitution on receptor affinity based on computer simulation of protein conformation and receptor molecule docking, explores the host adaptation evolution law of the virus in the transmission process, and provides an important scientific basis for early warning and prevention and control of a public health crisis possibly caused by the IDV.