38 results about "Coupling enzyme" patented technology

Isolated and / or recombinant enzymes that include surface binding domains, surfaces with active enzymes bound to them and methods of coupling enzymes to surfaces are provided. Enzymes can include large and / or multiple surface coupling domains for surface coupling.

The invention discloses a method for producing fermentable sugar through sulfonation, separation, coupling and enzymatic hydrolysis of cellulose compositions. The method comprises the following steps: firstly, cellulose raw materials are sliced or crushed and prepared by the pre-wet method; secondly, the cellulose raw materials are added into a water solution of sulfite and an alkaline compound, wherein the dosage of the sulfite is between 4 and 15 percent; the dosage of the alkaline compound is between 8 and 20 percent; the temperature of sulfonation reaction is between 120 and 180 DEG C; the reaction time, namely the heating-up time, is between 1 and 2 hours; and the soaking time is between 2 and 3.5 hours; and thirdly, cellulose products obtained through the sulfonation reaction which are not dissolved into a sulfonation reaction solution is dehydrated, cleaned and hydrolyzed in a cellulase reaction system which contains a buffer solution, and the fermentable sugar is obtained. The method can improve the yield of production of the fermentable sugar such as glucose, xylose and so on through transformation of the cellulose raw materials, improve the utilization rate of the raw materials, and achieve the aim of economy and high efficiency, and has important significance on promoting the commercial process of biomass energy - ethanol.

The invention provides a human proinsulin chemiluminescence immunoassay kit and a preparation method thereof. The kit provided by the invention comprises an anti-proinsulin reagent 1, an anti-proinsulin reagent 2, a magnetic separation reagent, a proinsulin calibrator, a chemiluminescence substrate, and a washing liquid. The invention also provides a preparation method of the kit. The preparation method comprises the following steps: coupling antibody and biotin, preparing the anti-proinsulin reagent 1, coupling enzyme and antibody, preparing the anti-proinsulin reagent 2, preparing the magnetic separation reagent, preparing the proinsulin correction sample, and preparing the chemiluminescence substrate. Compared to the proinsulin kit existing in the present market, the kit provided by the invention has the characteristics of high detection sensitivity, good repeatability, and low cost.

The invention relates to a method for extracting, separating and purifying four main anthocyanins from the blackcurrant residue. The object is to provide a simple, economic, green, eco-friendly, and high-yield method for extracting, separating and purifying four main high-purity anthocyanins from the blackcurrant residue. According to the method, the blackcurrant waste residue produced from industrial production is taken as a raw material, and homogenized pulverized coupling enzyme hydrolysis technology, negative-pressure cavitation reinforced extraction technology, macroporous adsorption resin concentration technology, medium-pressure quick-flash reverse chromatographic separation technology and low-temperature crystallization and recrystallization technology are adopted to obtain four anthocyanin monomer ingredients including delphinidin 3-O-glucoside, delphinidin 3-O-rutinoside, cyanidin 3-O-glucoside, and cyanidin 3-O-rutinoside and having the purity reach 95%. The method is abundant and available in raw materials, short in time consumption, less in solvent consumption, high in purity of obtained anthocyanins, low in production cost, and suitable for massive industrial production.

The invention discloses an ELISA (enzyme linked immunosorbent assay) kit for porcine circovirus type 2 (PVC2) antibody detection. The kit includes an antibody detection board, a sample diluting solution, an enzyme-labeled antigen, a washing solution, a substrate color developing solution, a stop solution, a positive serum and a negative serum. The Cap protein coated with antibody detection board of the kit is PCV2 eukaryotic expression nucleocapsid Cap protein, and the non-specific reactions are greatly reduced by using the Cap protein coupled enzyme-labeled antigen, so that the antibody detection accuracy is improved and the kit is more favorable for large-scale popularization and use.

The present invention relates to a composition comprising a coupled enzyme system for the rapid and efficient production of hydrogen peroxide by the coupling of a first enzyme system capable of hydrogen peroxide generation, to a second enzyme system which utilizes the non hydrogen peroxide product of the first enzyme system, and optionally is capable of generating further hydrogen peroxide.

A sterilization indicator system and method of using the system to determine efficacy of a sterilization process. The system includes a vial having a first compartment containing spores of one or more species of microorganism; a second compartment containing a growth medium with a disaccharide, an oligosaccharide or a polysaccharide in which the vial is free of monosaccharide; an enzyme, capable of acting upon the monosaccharide to yield reaction products and electron transfer, disposed on two or more electrodes adapted to carry an electrical signal resulting from the electron transfer, the pair of electrodes positioned to contact the combined contents of the first compartment and the second compartment during incubation; and an apparatus linked or linkable to the electrodes and adapted to detect and measure the electrical signal resulting from electron transfer when the enzyme acts upon the monosaccharide.

The invention relates to an enzymatic activity test method using combining integration and initial velocity method, which is characterized in that the classical initial velocity is measured when the concentration of initial substrate is 70% higher than that of the preset substrate, and the classical initial velocity is still within linearity range; when the substrate consumption proportion in 80% record period is higher than the lower limit required in integration, the fitting reaction curve of integral velocity equation is used to determine the maximum reaction velocity using reaction time as independent variable, then the maximum reaction velocity is calculated as the initial velocity with preset substrate concentration is 93%; meanwhile, the slope of classical initial velocity and computation initial velocity to the enzyme amount reaction curve; when the substrate consumption proportion of the coupling enzyme reaction system in 80% record period is higher than the lower limit required in integration, the fitting enzyme reaction curve is used to determine the initial velocity using corresponding numerical integral velocity equation, otherwise the classical initial velocity is directly measured; total time for the optimization monitoring reaction guarantees the combination of the two methods.

The invention discloses a method catalytically synthesizing (S)-N, N-dimethyl-3-hydroxy-(2-thiofuran)-1-propylamine((S)-DHTP) by aldehyde ketone reductase recombinant strain crude enzyme system, and belongs to the technical field of biological catalysis asymmetric conversion. An auxiliary substrate and original bulk coenzyme NADP<+> are added in the recombinant strain crude enzyme system to accelerate regeneration cycle of coenzyme NADPH in the system; the reaction substrate is N, N-dimethyl-3-ketone-(2-thiophene)-1-propylamine (DTKP); the recombinant strain is E. coliBL21(DE3)(pETCPAR4); aldehyde ketone reducing enzyme gene cpar4 comes from Candida parapsilosis; the gene coded aldehyde ketone reducing enzyme gene CPAR4 catalytically and asymmetrically reduces the DKTP into (S)-DHTP. The method utilizes the cell-free system to conduct catalytic reaction; coupling enzyme required by regeneration of the coenzyme is not extra added in the reaction system; direct acting efficiency of the enzyme and the substrate is improved; reaction time is shortened; the conversion effect is relatively good.

The invention discloses a method for preparing (R)-phenylglycol from SD-AS sequence coupled (R)-carbonyl reductase and glucose dehydrogenase and belongs to the technical field of biological catalytic asymmetric transformation. The invention provides recombinant escherichia coli E. coli RIL / pET-R-SD-AS-G. The recombinant escherichia coli E. coli RIL / pET-R-SD-AS-G is preserved in the China center for type culture collection (CCTCC) and has a preservation number of CCTCC NO: M2015170. R-carbonyl reductase and glucose dehydrogenase are coupled by a coexpression way, and the coupled enzyme, a cultured recombinant strain and 2-hydroxyacetophenone as a substrate undergo a catalytic asymmetric transformation reaction under optimized biotransformation reaction conditions to produce (R)-phenylglycol with optical purity of 100% and a yield of 99.9%. The method utilizes a SD-AS sequence high efficiency coupling double-enzyme technology, solves the problem of limitation of chiral catalytic reaction coenzyme regeneration cycle and provides an effective approach for high efficiency bio-preparation of the (R)-phenylglycol.

The present invention relates to a composition comprising a coupled enzyme system for the rapid and efficient production of hydrogen peroxide by the coupling of a first enzyme system capable of hydrogen peroxide generation, to a second enzyme system which utilizes the non hydrogen peroxide product of the first enzyme system, and optionally is capable of generating further hydrogen peroxide.

A method for producing chitosan oligosaccharides coupled with enzymatic degradation of chitosan and membrane separation is characterized in that the production process is as follows: (1) preparing chitosan oligosaccharides by enzymatic degradation of chitosan; (2) adopting hollow fiber or flat ultra- Combine the filter with the degradation reaction to separate the active oligosaccharides in situ; (3) filter the ultrafiltrate with a nanofilter to separate high-concentration active chitosan oligosaccharides. The invention realizes the integration of reaction and separation, and prevents secondary degradation of active oligosaccharides. Since the process realizes continuous operation, the degrading enzyme can be used continuously, the yield is improved, the cost is reduced, and it is beneficial to realize industrialization.

An economical and expedient method is disclosed for the preparation of alpha-hydroxy-carboxylic acids or salts thereof in very high enantiomeric purity which comprises oxidizing a corresponding alpha-amino-carboxylic acid or salt thereof using an amino acid deaminase followed by reducing the corresponding alpha-keto-carboxylic acid or salt produced using a D- or L-lactate dehydrogenase in the combination with an electron donor and an enzyme / substrate system for recycling the electron donor. The resulting alpha-hydroxy-carboxylic acids, hydrates, and salts thereof are valuable components and intermediates in the preparation of chiral compounds, especially pharmaceuticals. This invention also relates to the use of alpha-amino-carboxylic acids, hydrates, and salts thereof and a coupled enzyme system in the production of alpha-hydroxy-carboxylic acids, hydrates, and salts thereof.

The present invention relates generally to enzyme assays and more particularly to a rapid, sensitive, reliable and robust homogeneous assay for acetyl CoA carboxylase activity comprising a coupled enzyme assay in a scintillation proximity format suitable for high-throughput screening. In one aspect, the assay couples ACC and FAS and the product, palmitic acid is then detected by scintillation counting. The invention also relates to the identification of modulators of ACC activity.

The invention discloses a liquid stable enzymic method homocysteine kit. The liquid stable enzymic method homocysteine kit is characterized by comprising (1) a stable substrate system and (2) stable S-adenosyl-L-homocysteine hydrolase, wherein the stable substrate system comprises an NADH circulatory system and an S-adenosyl methionine polyanion system, and the stable S-adenosyl-L-homocysteine hydrolase improves the overall stability of the kit. By the adoption of the liquid stable enzymic method homocysteine kit, a stable system of S-adenosyl methionine and reduced coenzyme I is provided, the patented technology of enzyme is introduced so that the overall stability of the kit can be improved, and therefore the demand of clinical examination for the liquid stable enzymic method homocysteine kit is met; in addition, the defect that in the prior art, the clinical application of a kit is limited due to the fact that the stability of the substrate S-adenosyl methionine (SAM) is poor and the thermal stability of an adopted coupling enzyme is poor is overcome.

Bertrand(1982) hydrolytic generates trophicardyl from trophicardyl acid by nucleotidase, and gains H2O2 through the reaction of the enzyme1 and enzyme2. Use of H2O2 from Trinder's reaction to combine hydrogen provider3,5-double CL-2-hydroxide benzene sulphur acid and 4- to create a amaranth in the situation of the catalase. By watching the velocity of increase of the degree of the amaranth absorbing light at 510 nms to gain 5' nucleotidase activity. The invention uses the aniline compound ADOS, ADPS, ALPS, TODB, TOOS and TOPS as hydrogen provider for Trinder's reaction and increases the reaction ability. The invention also involves a reagent box used for implementing the method mentioned above.

The present invention relates to coupled enzyme assays. In particular, the present invention provides a coupled fluorescent assay for detection of S-adenosylmethionine (AdoMet)-dependent methyltransferase activity.

The invention discloses a method and a device for producing fuel ethanol by solid state fermentation of fiber raw materials. The method comprises the following steps of: coupling enzyme generated by the solid state fermentation of mixed bacteria with ethanol generated by the synchronous fermentation of a substrate, and continuously carrying out enzymolysis on the substrate sufficiently by the effective enzyme synthesized by the microorganism and the metabolism of the microorganism in the process of solid state fermentation to promote cellulose and hemicellulose in the substrate to be effectively degraded; synchronously saccharifying and fermenting a carbon resource matrix to be converted into ethanol sufficiently by a mixture of the high-concentration sugar generated by degrading the substrate, the un-degraded matrix of the sugar and the compound enzyme synchronously synthesized by means of fermentation, wherein the ethanol yield is more than 37% of the theoretical conversion rate. The application of high-cost commercial cellulose can be avoided in the process, and the production cost of the fuel ethanol can be greatly reduced. The solid state tray fermentation device provided by the invention is simple in structure, and easy to manufacture, and the defect of enzyme production by fermenting single bacterial strain and the problems that the matrix in a solid state fermentation reactor is uneven in humiture distribution and is easily infected with infectious microbe and the like can be overcome.

The invention relates to an EV71 single-stranded DNA aptamer and a chemiluminescent detection kit for detecting an enterovirus type-71 by utilizing double aptamers. The aptamer is an aptamer V11 having a sequence shown as SEQ ID NO.1, an aptamer V21 having a sequence shown as SEQ ID NO.2, and an aptamer V7 having a sequence shown as SEQ ID NO.3. The kit takes the aptamer V21 as a capture probe andthe aptamer V11 as a signal probe. The chemiluminescent detection kit specifically comprises capture probe coupled immunomagnetic beads, an assay buffer solution, a signal probe coupled enzyme conjugate, an SA-HRP solution and a luminescent substrate solution. The kit has high specificity for EV71 detection and can qualitatively detect EV71 from a cerebrospinal fluid, a herpes fluid, a pharyngealswab, an anal swab and other samples of a suspected hand-foot-mouth disease patient. By utilizing the kit, detection operation is simple and rapid, and the detection process only needs about 45 minutes.

The invention discloses a quantitative determination method of cardiac myosin binding protein C. According to the method, firstly, antibody coupling super paramagnetic micro particles and samples to be tested take specific combination reaction to obtain a first reactant, wherein the antibody coupling super paramagnetic micro particles are products obtained by coupling super paramagnetic micro particles and anti-cMyBP-C monoclonal antibodies A; then, the first reactant is taken to perform specific combination reaction with enzyme-labeled articles to obtain a second reactant, wherein the enzyme-labeled articles are products obtained by coupling enzymes and anti-cMyBP-C monoclonal antibodies B; finally, chemiluminescence substrates and the second reactant are taken to take enzymatic reaction;luminous signals are determined; the concentration value of cMyBP-C is obtained. The technology mode of the invention is a sandwich method. Compared with a cMyBP-C ELISA kit, the detection method andthe prepared kit have the advantages that the detection sensitivity and the linear range can be greatly improved. The technology evaluation on the prepared kit shows that the analysis sensitivity is0.05 ng / ml; the detection range is 0.1 to 50 ng / ml; high correlation and coincidence rate to clinic AMI patients are high.

The present invention relates to coupled enzyme assays. In particular, the present invention provides a coupled fluorescent assay for detection of S-adenosylmethionine (AdoMet)-dependent methyltransferase activity and S-adenosylhomocysteine hydrolase enzyme activity.

The invention discloses a method for preparing pharmaceutical dextran and levulose by bi-enzyme immobilization coupled multistage membrane separation, which comprises the following steps: co-immobilizing dextran sucrase and dextranase in sodium alginate microspheres to obtain bi-enzyme-immobilized spheres; adding the bi-enzyme-immobilized spheres into a sucrose solution used as a substrate to form an immobilized bi-enzyme / sucrose reaction system; and after the reaction product is separated by a multistage membrane system composed of hyperfiltration membranes and a nanofiltration membranes, directly obtaining the pharmaceutical dextran in the hyperfiltration membrane separation stage, and obtaining the high-purity levulose solution in the last nanofiltration membrane separation stage, wherein the enzymes and macromolecular dextran contained in the trapped solution of the first-stage hyperfiltration membrane return to the immobilized bi-enzyme / sucrose reaction system to participate in the reaction again, thereby implementing cyclic utilization of the enzymes. The invention has the advantages of lower cost, environment friendliness, high safety and high efficiency, is simple to operate, implements oriented adjustable production of high-quality pharmaceutical dextran with target molecular weight, and obtains the high-purity high-added-value levulose byproduct.

The invention discloses a high-specificity coupling enzyme YND mutant of 3', 5'-adenosine diphosphate based on molecular docking, the gene sequence of the YND mutant is SEQ ID NO.2, SEQ ID NO.3 or SEQ ID NO.4, and the amino acid sequence of the YND mutant is SEQ ID NO.6, SEQ ID NO.7 or SEQ ID NO.8. The invention designs and modifies a coupling enzyme with strong PAP specificity by taking yeast 3 ', 5'-adenosine diphosphate enzyme with a known crystal structure as a starting point. The modified enzyme is subjected to inducible expression by an escherichia coli expression system and stored in a storage solution, which can maintain activity for 3 months at-80 DEG C, modified enzyme is used by being diluted with a corresponding buffer system, the use is convenient and the reaction system can be replaced, and the problem of limitation of a system buffer substance on ST activity detection is solved.

Aldehyde and ketone reductase crude enzyme system catalytic synthesis of recombinant bacteria ( S )‑N,N‑Dimethyl‑3‑hydroxy‑3‑(2‑thiophene)‑1‑propylamine (( S )‑DHTP), which belongs to the technical field of biocatalytic asymmetric transformation. The present invention is to add auxiliary substrate and initial amount of coenzyme NADP in recombinant bacterial crude enzyme system + Promote the regeneration cycle of the coenzyme NADPH in the system, the reaction substrate is N,N-dimethyl-3-ketone-3-(2-thiophene)-1-propanamine (DTKP), and the recombinant bacteria express the aldehyde and ketone reductase gene of E. coli BL21(DE3)(pETCPAR4), aldehyde and ketone reductase gene cpar4 from Candida parapsilosis ( Candida parapsilosis ) CCTCC NO: M 203011, the gene encodes aldehyde and ketone reductase CPAR4, which catalyzes the asymmetric reduction of DKTP to ( S )‑DHTP. The invention utilizes the cell-free system to catalyze the reaction without additionally adding coupling enzymes required for coenzyme regeneration in the reaction system, improves the efficiency of the direct action of the enzyme and the substrate, shortens the reaction time, and obtains better conversion effects.

A sterilization indicator system and method of using the system to determine efficacy of a sterilization process. The system includes a vial having a first compartment containing spores of one or more species of microorganism; a second compartment containing a growth medium with a disaccharide, an oligosaccharide or a polysaccharide in which the vial is free of monosaccharide; an enzyme, capable of acting upon the monosaccharide to yield reaction products and electron transfer, disposed on two or more electrodes adapted to carry an electrical signal resulting from the electron transfer, the pair of electrodes positioned to contact the combined contents of the first compartment and the second compartment during incubation; and an apparatus linked or linkable to the electrodes and adapted to detect and measure the electrical signal resulting from electron transfer when the enzyme acts upon the monosaccharide.

The invention discloses a device and method for gradient concentration by a coupling enzyme-membrane method using residual heat for mechanical supercharging. The device comprises an ultra filtration membrane filtering unit, a reverse osmosis membrane filtering unit and a mechanical supercharging concentrating unit, wherein the ultra filtration membrane filtering unit comprises a balancing barrel, a first enzymatic hydrolysis tank and an original ultra filtration membrane which communicate each other through a pipeline. The device disclosed by the invention can concentrate raw material soup under low energy consumption level by level so as to obtain a concentrated solution of different concentrations; the first enzymatic hydrolysis tank is additionally arranged in the ultra filtration membrane filtering unit, so that protein in the raw material soup can be separated through membranes with different bore diameters so as to obtain soup in a target molecular weight segment, and the characteristic attributes of products are improved; mechanical supercharging concentration is performed, so that the material soup with a high concentration is obtained. Through the device and the application method disclosed by the invention, the target concentrated solution with different concentrations and different molecular weights can be obtained, the original flavor of materials is well maintained, a great number of energy resources are saved, and the method and the method are particularly suitable for concentration of bone soup.

The invention discloses a high-efficiency polypeptide-polypeptide coupling system and method based on a disordered protein coupling enzyme. By designing and splitting the protein domain CnaB2 and optimizing it, a three-component system is obtained: spy tag (SpyTag), BD tag (BDTag) and spy stapler enzyme (SpyStapler), so that SpyStapler can catalyze the coupling between SpyTag and BDTag Linkage reaction, and prepare different fusion proteins containing SpyTag and BDTag two tags and carry out coupling or cyclization reaction. Based on the polypeptide-polypeptide coupling system of the present invention, an active protein with high purity and stable function can be obtained by means of gene coding; the spy stapler enzyme coupling spy label and BD label have high reaction efficiency, and some of the staples are stapled after the reaction The mechanical enzyme falls off, and the coupling part of the obtained cyclization product has a smaller molecular weight. The protein tag of the present invention is applicable to various functional proteins and can be fused and expressed at any position of the protein.

The present invention relates to genetically modified enzymes obtained by rational design of the active site binding pocket of the prototypic enzyme 4-hydroxyphenylacetate 3-hydroxylase (4HPA3H) for hydroxylating a 4-hydroxyphenyl compound to yield a 3,4-dihydroxyphenyl compound and to biotechnological methods including in vivo and in vitro methods using said enzymes or catalytically active fragments thereof. Further provided is a method either using a suitable oxidase or hydroxylase further enabling the subsequent site specific methylation of the 3,4-dihydroxyphenyl compound in a coupled enzymatic reaction by providing a suitable O-methyltransferase. Finally, compositions obtainable by the aforementioned methods are disclosed.