120 results about "Specific activity" patented technology

The invention discloses an L-aspartic acid transaminase mutant and application thereof. The L-aspartic acid transaminase mutant is obtained by performing single mutation on the 104th amino acid of the amino acid sequence of L-aspartic acid transaminase. The L-aspartic acid transaminase mutant is used for synthesizing an L-phenylglycine derivative. The L-aspartic acid transaminase mutant is from escherichia coli, homologous sequence analysis and molecular docking simulation are carried out, site-specific mutagenesis verification is combined, and single-point mutation is carried out on the 104 amino acid to obtain the T104S mutant. The L-aspartic acid transaminase mutant disclosed by the invention can be used for efficiently catalyzing to generate an L-phenylglycine derivative, and compared with a wild type, the specific activity of the T104S mutant to o-chlorophenoxyacetic acid is 3.2 times that of the wild type.

The invention provides an endoglucanase mutant and application of the endoglucanase mutant. The endoglucanase mutant comprises: 1) a protein which has at least one of the following sites in SEQ ID NO: 29: D194, T232 or G247, has a mutated amino acid sequence, and has endoglucanase activity; or 2) a protein which has an amino acid sequence with homology of at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 99% with the protein in 1) and has endoglucanase activity. Compared with a wild type, the endoglucanase mutant has higher specific activity and is beneficial to wide application of beta-endoglucanase in industry.

The invention discloses a xylanase mutant with synchronously improved thermal stability and pH tolerance and application of the xylanase mutant, and belongs to the field of gene engineering and protein engineering. According to the invention, xylanase AbXyL10A from Agaricus bisporus var burnettii is taken as a female parent, and two amino acid sites of Thr234 and Arg268 are subjected to mutation, so that two types of xylanase mutants T234N and R268L are obtained. Compared with wild type xylanase, the thermal stability and pH tolerance of the two xylanase mutants are obviously improved. In the aspect of specific activity, when the beech xylan is used as a substrate, the specific activity of the two xylanase mutants is obviously improved compared with that of wild type xylanase. The invention provides a theoretical reference for the improvement of xylanase, and the obtained xylanase mutant with synchronously improved thermal stability and pH tolerance has a huge application prospect in the aspects of animal feed and biomass degradation sugar production.

The invention belongs to the technical field of biology, and discloses a UDP-glucosyltransferase mutant and an application thereof, and a preparation method of stevioside rebaudioside D. The UDP-glucosyltransferase mutant comprises a UDP-glucosyltransferase mutant, compared with the original UDP-glucosyltransferase comprising an amino acid fragment with the sequence shown as SEQ ID NO: 1, the UDP-glucosyltransferase mutant provided by the invention comprises H18P mutation and / or R370H mutation, and the mutation endows the UDP-glucosyltransferase mutant with higher specific activity for catalyzing the conversion of rebaudioside A into rebaudioside D, so that the UDP-glucosyltransferase mutant can be used for catalyzing the conversion of rebaudioside A into rebaudioside D; the high-efficiency and sufficient conversion from the rebaudioside A to the rebaudioside D can be realized, the realization of large-scale and high-efficiency production of the rebaudioside D is facilitated, and the method has a good application prospect.

The present application relates to the field of genetic engineering and PET material degradation, and particularly relates to a PET degrading enzyme mutant and its application in PET material degradation. The present application improves the activity and heat resistance of PET degrading enzyme MuPETase derived from Mumia sp., and screens to obtain mutants Mut1-3, Mut5-8 which show good degradation capacity to PET material under acidic conditions, and the amino acid sequences are shown in SEQ ID NO. 3-5, SEQ ID NO. 7-10 respectively. Compared with the original PET degrading enzyme, the heat stability and / or specific activity of the obtained mutant are significantly improved, and the degradation effect on PET material is greatly improved. The obtained mutant has important application value in the biological degradation treatment of single and mixed PET material waste.

This invention discloses a lipase mutant, an immobilized enzyme, and their applications, belonging to the field of bioengineering. The SpL enzyme mutant K193R / I134V described in this invention exhibits a specific activity of 7-ACA that is 1.6 times higher than before the mutation, and its half-life at 45℃, 50℃, and 55℃ is increased by 1.9 times, 2.6 times, and 3.8 times, respectively. Immobilizing the his-tagged SpL enzyme on a hydrophobically modified nickel-doped SBA-15 mesoporous molecular sieve preserves the enzyme's activity and improves its thermal stability and reusability. Using this immobilized enzyme to catalyze the production of D-7-ACA allows for at least 10 reusable batches, maintaining a conversion rate above 99%.

Disclosed herein are compositions comprising recombinant acid sphingomyelinase (rASM) having a desired purity, specific activity and / or rASM isoform. Methods for producing and purifying such compositions, including chromatography steps, are also provided. Additionally, methods for modulating rASM specific activity in a composition, and methods for modulating rASM isoform in a composition are provided. The methods disclosed herein may be particularly useful for the preparation of pharmaceutical compositions comprising rASM for treating acid sphingomyelinase deficiency (ASMD).

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 present invention discloses an engineered protein glutaminase mutant and its method for active secretion expression in Pichia pastoris. The present invention successfully improves the catalytic activity of FBPG through surface charge modification, and the specific activity of mutant F3 reaches 9.43 times that of the wild type. The secreted expression of FBPG is achieved in Pichia pastoris, and under the regulation of the P DH promoter, the active expression of FBPG is achieved by introducing the Kex2 cleavage site. After overexpressing the transcription factor Sko1, the yield of active FBPG reaches 2610 U / L in a 7 L bioreactor, providing a feasible technical solution for the application of FBPG in the food industry, and also providing new ideas and methods for the active expression of similar enzymes.

The invention discloses a method for preparing < 18 > F-m-fluorobenzylguanidine, and solves the technical problems of complex reaction steps, low radiochemical yield and low specific activity in a method for synthesizing < 18 > F-mFBG. The invention comprises a phenolic ether precursor compound of a core skeleton corresponding to 18F-mFBG, a synthesis method of the phenolic ether precursor compound, and a method for realizing F-18 labeling and preparation of mFBG by using the precursor compound. Specifically, 18F-mFBG labeling and preparation are realized by constructing a key phenolic ether precursor compound and performing simple and efficient photocatalytic deoxidation fluorination reaction on aromatic hydrocarbon. The method has the advantages of few steps, high yield, simplicity and convenience in operation, good repeatability and good automatic production potential. The radioactive tracer 18F-mFBG synthesized by the method is high in specific activity, chemical purity and radioactive purity, and has important value for clinical popularization of targeted NET molecular function PET imaging.

The invention relates to an anti-mouse IgG nano antibody and alkaline phosphatase fusion protein as well as a preparation method and application thereof. The anti-mouse IgG nano antibody and alkaline phosphatase fusion protein sequentially comprises an anti-mouse IgG nano antibody, a connecting peptide and shewanella alkaline phosphatase from an amino terminal to a carboxyl terminal. The fusion expression of the anti-mouse IgG nano antibody and shewanella alkaline phosphatase in escherichia coli is designed, so that various defects of a chemical labeling method can be avoided, a high-purity and high-specific activity enzyme-labeled antibody can be prepared, the preparation period of a second antibody can be shortened, the preparation process of the second antibody can be simplified, the stability of batches can be improved, and the dependence on experimental animals can be reduced. The prepared anti-mouse IgG nano antibody and alkaline phosphatase fusion protein can be applied to the field of immunodetection, such as a chemiluminescence immunoassay method, an enzyme-linked immunosorbent assay method, an enzymatic fluorescence immunoassay method, an immunoblotting method and the like.

The invention provides an N-deacetylase / N-sulfotransferase and an application of the N-deacetylase / N-sulfotransferase. According to the invention, systematic stability design and surface electrostatic optimization are carried out on the N-deacetylation structural domain of NDST, and multiple engineering strategies such as structural domain fusion and linker peptide optimization are combined to synergistically improve the specific activity, substrate affinity and bifunctional catalytic synergy of enzyme. Experimental results show that the specific enzyme activity of the modified NDST mutant is obviously higher than that of natural enzyme while the difunctional catalytic capability is maintained.

The invention discloses a preparation method of pichia pastoris expression secreting type deglycosylation alkaline phosphatase, and belongs to the technical field of biological engineering. According to the invention, the GS115-BIAP II pichia pastoris genetically engineered bacterium capable of secreting and expressing bovine intestine alkaline phosphatase BIAP II is prepared by using a conventional method. The method comprises the following steps: carrying out site-directed knockout on an ALG3 gene for coding alpha-1, 3-mannosyl transferase and an OCH1 gene for coding alpha-1, 6-mannosyl transferase by using a CRISPR-Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats-associated Protein 9) technology to prepare a glycosylation gene defect type engineering bacterium GS115-delta ALG3-delta OCH1-BIAP II. The secretory expression BIAP II is deglycosylated alkaline phosphatase, and the specific activity of the secretory expression BIAP II is close to that of natural BIAP II. The deglycosylated alkaline phosphatase can improve the antibody coupling efficiency and marking uniformity under specific conditions. The method has remarkable advantages in chemiluminescence diagnosis, high-sensitivity immunodetection and development of multifunctional biological coupling reagents.

The invention relates to the technical field of gene engineering and protein modification, in particular to a high-specific-activity cellulase mutant and application thereof. Compared with the wild type cellulase, the specific activity of the mutants respectively containing single mutation sites of P14A, P19T, S75A, S109N, H118P, M123I, M123L, I130L, I130Q, Y167E, Y167F, Y167W, A176S, A176F, D177S and L192I, provided by the invention, is generally improved by 10.4%-76.1% under a neutral condition. Wherein the specific activity of a single-site mutant containing I130L is the highest, and reaches 183.64 U / mg. The production cost of the cellulase mutant is remarkably reduced, so that wide application of the cellulase mutant is promoted.

The invention relates to the technical field of enzyme engineering, and particularly discloses a phytase mutant and application thereof. The mutant T273K with improved heat resistance is obtained through rational design and site-directed mutagenesis screening on the basis of aspergillus tubingensis phytase. The thermal stability and catalytic ability of the mutant are obviously improved compared with those before mutation; after heat preservation is carried out for 90 min at the temperature of 60 DEG C, the relative enzyme activity is 69.4%, and compared with WT, the relative enzyme activity is improved by 20.4%; the specific activity is improved by about 77.78% compared with that of WT, and the catalytic efficiency kcat / Km is improved by 50.56% compared with that of WT; and the enzyme activity retention rate reaches 33.8% under the condition of 10% salinity. The phytase mutant T273K has the comprehensive characteristics of salt resistance, heat resistance and the like, the phytase mutant T273K can better exert the phytic acid enzymolysis effect in the food processing process, turbidity generated by phytic acid in liquid food such as soy sauce, vinegar, cooking wine and the like can be remarkably reduced, so that the product quality is improved, and the phytase mutant T273K has a wide application prospect.

The invention discloses pseudomonas aeruginosa 24F6 and application thereof, and belongs to the technical field of microorganisms. The invention discloses a pseudomonas aeruginosa strain 24F6 with the preservation number of CGMCC No.35032. The chicken feather degradation rate is 86%, and the wool degradation rate is 40%. The specific activity of the strain cultured for 24 hours under the conditions that the pH value is 7.0 and the temperature is 40 DEG C is 119.67 U / mg. The invention also relates to application of the keratin degrading agent in preparation of keratin degrading products. The pseudomonas aeruginosa 24F6 has the following advantages that a plurality of bacteria are separated from the excrement of the larva of the costustomoth, strains with different keratin degradation capacities are screened out through a keratin selective culture medium, and the pseudomonas aeruginosa 24F6 is a strain which is reported for the first time and has unique keratin degradation characteristics; the specific activity of the strain under the optimal fermentation condition is obviously higher than that of reported similar strains, the degradation rate of natural chicken feathers is as high as 86%, and the strain shows good industrial application potential.

The present invention discloses a cyanase mutant, nucleic acid, recombinant expression vector and their applications, belonging to the field of bioengineering. The cyanase gene derived from Escherichia coli E. coli K-12 substr. MG1655 was subjected to site-directed mutagenesis, so that the encoded amino acid sequence was mutated from glycine G to alanine A at the 37th position, from lysine K to arginine R at the 154th position, and from glutamine Q to histidine H at the 51st position. The specific activity of the cyanase mutant G37A / K154R / Q51H of the present invention against potassium cyanate is 2.4 times that before mutation, and the half-lives at 30 °C and 40 °C are increased by 2.1 times and 2.6 times respectively. The cyanase parent of the present invention cannot catalyze potassium cyanide, while the specific activity of the mutant G37A / K154R / Q51H against potassium cyanide reaches 5.5 U / mg, and the half-lives at 30 °C and 40 °C are 2.5 h and 3.0 h respectively. The cyanase mutant of the present invention can efficiently catalyze the decomposition of the substrates potassium cyanate and potassium cyanide, so as to achieve the purpose of degrading cyanide-containing wastewater, and can be widely applied to industries such as chemical engineering, pharmaceuticals and metallurgy, and has broad application prospects.

The invention belongs to the technical field of genetic engineering and protein engineering modification, and particularly relates to a high-specific-activity cellulase mutant and application thereof. Compared with wild cellulase, the specific activity of the mutant is improved by 51.9% and reaches 319 U / mg, and an unexpected technical effect is achieved. The mutant can be widely applied to the field of textile industry, is uniform in flower formation, small in flower point and strong in stereoscopic impression, most importantly, the strength loss of the fabric is very small, the strength retention rate is improved by 10-15% compared with that of the fabric treated by wild type cellulase, the use amount of the cellulase can be greatly reduced, the cost is reduced, and the application prospect is wide.

Disclosed is a method for producing hyaluronidase or a variant thereof. Specifically, the method is capable of changing the N-glycan levels under culture conditions including a controlled concentration of glucose in the culture medium and a decreased culture temperature for a specific culture time period, thereby increasing the specific activity by 10% or more and improving the quality and production yield.

Provided are high-specific-activity cellulase mutants and use thereof. Compared to the wild-type cellulase, the specific activity of the provided mutants that comprise single mutation sites P14A, P19T, S75A, S109N, H118P, M123I, M123L, I130L, I130Q, Y167E, Y167F, Y167W, A176S, A176F, D177S, and L192I, respectively, under neutral conditions is generally increased by 10.4% to 76.1%. The single-point mutant comprising I130L has the highest specific activity, which reaches 183.64 U / mg. The production costs of the cellulase mutants are significantly lower, which facilitates the wide use thereof.

The application discloses a xylanase mutant with simultaneously improved thermal stability and pH tolerance and application thereof, and belongs to the field of genetic engineering and protein engineering. The xylanase AbXyL10A from Agaricus bisporus var burnettii is used as a parent to obtain two xylanase mutants T234N and R268L after mutation of two amino acid sites Thr234 and Arg268. Compared with the wild-type xylanase, the thermal stability and pH tolerance of the two xylanase mutants are obviously improved. In terms of specific activity, when beech xylan is used as a substrate, the specific activity of the two xylanase mutants is obviously improved compared with the wild-type xylanase. The application provides a theoretical reference for improvement of xylanase, and the obtained xylanase mutant with simultaneously improved thermal stability and pH tolerance has great application prospect in animal feed and biomass degradation sugar.

The invention discloses a method for detecting the specific activity of a recombinant pseudomonas aeruginosa vaccine rPO stock solution. The method comprises the following steps: (1) coating an elisa plate by using OprI-B052 as a capture antibody; (2) adding a sample to be detected into the coated ELISA plate; (3) diluting the enzyme-labeled antibody PcrV-A039, then carrying out a chromogenic reaction, and measuring the OD450 / 630 nm value; and (4) preparing a standard curve by using the rPO reference, and calculating the specific activity of the rPO antigen in the detected sample according to the standard curve. According to the invention, OprI-B052 is used as a coating antibody, and paired PcrV-A039 is used as a detection antibody. The specific activity of the recombinant pseudomonas aeruginosa vaccine rPO stock solution can be quantitatively detected, and the method has good stability and repeatability.

The invention provides a 5-hydroxymethylfurfural oxidase mutant and application thereof, and belongs to the technical field of biology. The sequence of the 5-hydroxymethylfurfural oxidase mutant is as shown in SEQ ID NO: 4. The invention also provides a gene for coding the 5-hydroxymethylfurfural oxidase mutant, a recombinant vector and a recombinant cell containing the gene, and applications of the 5-hydroxymethylfurfural oxidase mutant, the recombinant vector and the recombinant cell in catalyzing 5-hydroxymethylfurfural to generate 2, 5-furandicarboxylic acid. According to the 5-hydroxymethylfurfural oxidase mutant provided by the invention, the catalytic reaction effect is obviously superior to that of a wild type, the specific activity is improved to 21.4 times, and the FDCA yield is improved to 18.0 times. The mutant can tolerate high substrate concentration, the reaction product is single and high in purity, the FDCA yield is larger than 95%, and the mutant has high industrial value and application prospects.

This invention relates to a glutathione S-transferase derived from Antarctic krill, its encoding gene, and its applications, belonging to the field of enzymology. The amino acid sequence of the glutathione S-transferase is shown in SEQ ID NO.1, and the amino acid sequence of the gene encoding the glutathione S-transferase is shown in SEQ ID NO.2. The specific activity of the glutathione S-transferase of this invention can reach up to 6.01 μmol / min / mg. The optimal temperature and optimal pH are 20 °C and 8, respectively. The Antarctic krill-derived glutathione S-transferase of this invention exhibits good catalytic activity and substrate affinity at relatively low temperatures.

This invention provides a method for purifying high-purity recombinant follicle-stimulating hormone (FSH). Specifically, it includes the following steps: (a) providing a raw material solution containing recombinant FSH; (b) sequentially subjecting the raw material solution to: (b1) ammonium sulfate precipitation; (b2) hydrophobic interaction chromatography (HIC); (b3) low pH incubation to inactivate the virus; and (b4) reversed-phase chromatography (RPC); wherein steps (b1), (b2), (b3), and (b4) cannot be performed in any arbitrary order. Compared with existing preparation methods, the recombinant FSH obtained by the method of this invention is simpler and easier to operate, and the purity of the recombinant FSH obtained by the method of this invention can reach up to 99.5% w / w, with a specific activity as high as 16000 IU / mg.

The application discloses a method for detecting specific activity of recombinant Pseudomonas aeruginosa vaccine rPO stock solution, and comprises the following steps: (1) using OprI-B052 as a capture antibody to coat an enzyme-labeled plate; (2) adding a sample to be detected into the coated enzyme-labeled plate; (3) diluting an enzyme-labeled antibody PcrV-A039 and then performing a color reaction to measure an OD 450 / 630 nm value; and (4) preparing a standard curve by using a rPO reference product, and calculating the specific activity of the rPO antigen in the sample according to the standard curve. In the application, OprI-B052 is used as a coating antibody, and PcrV-A039 is used as a detection antibody. The specific activity of the recombinant Pseudomonas aeruginosa vaccine rPO stock solution can be quantitatively detected, and the application has good stability and repeatability.

The invention relates to the technical field of nuclear material analysis, in particular to a plutonium solution specific activity determination method and application, and the plutonium solution specific activity determination method comprises the following steps: taking a plutonium solution with known specific activity A1 as a standard solution, determining the liquid flash counting rate of the standard solution, and recording the liquid flash counting rate as C1; obtaining a second-order conductance spectrum of the standard solution by using an ultraviolet-visible absorption spectrum method, determining the peak area of a characteristic absorption peak of tetravalent plutonium in the second-order conductance spectrum of the standard solution, and recording the peak area as S1; measuring the liquid flash counting rate of the to-be-measured solution, and recording the liquid flash counting rate as C2; obtaining a second-order conductance spectrum of the to-be-detected solution by using an ultraviolet-visible absorption spectrometry, determining the peak area of a characteristic absorption peak of tetravalent plutonium in the second-order conductance spectrum of the to-be-detected solution, and recording the peak area as S2; and calculating the specific activity A2 of the solution to be detected according to the following formula. According to the method, liquid scintillation counting and ultraviolet-visible spectrophotometry are combined, so that combination of activity response and concentration response is realized, and acquisition of isotope composition of a plutonium solution to be detected and complex sample pretreatment are not needed.

An anti-fibroblast activation protein alpha (FAP) polypeptide comprising at least one single-domain antibody directed against FAP. A pharmaceutical composition comprising an anti-FAP single-domain antibody conjugated to a radioisotope at a ratio that maximizes specific activity but does not compromise affinity. A method of reducing a tumor is a subject in need thereof, comprising administering the pharmaceutical composition to a subject.

The present invention relates to the technical fields of genetic engineering and protein engineering modification, and particularly relates to a high-specific-activity alginate lyase mutant and its application. The alginate lyase mutant provided by the present invention has a significantly higher specific activity than the wild-type alginate lyase under the low-temperature condition of 40°C, achieving an unexpected technical effect. It can be widely used in the field of seaweed processing, effectively improving the enzymatic hydrolysis efficiency of seaweeds such as Sargassum and Ascophyllum nodosum, and having a broad market prospect.

The invention discloses a restriction enzyme XcmI mutant capable of improving activity and stability and application of the restriction enzyme XcmI mutant, and belongs to the technical field of biology, and the amino acid sequence of the restriction enzyme XcmI mutant is shown as SEQ ID NO.1. The specific activity of the XcmI mutant is more than three times that of a wild type, the mutant can still remain 100% of activity after being stored at 37 DEG C for 48 hours, while only 10% of activity of the wild type remains, and after being treated by 20mM H2O2 for 30 minutes, the mutant can still remain about 30% of activity while only 10% of activity of the wild type remains. Compared with a wild type, the restriction enzyme XcmI mutant disclosed by the invention has the advantages that the activity, the thermal stability and the oxidation stability are remarkably improved, and the restriction enzyme XcmI mutant has a relatively good application value.