47 results about "ATP regeneration" patented technology

The invention discloses a method for regenerating ATP by enzymatic method, which comprises the following steps: (1) preparing and obtaining ATP regenerating enzyme; (2) regenerating ATP in reaction liquid; (3) separating products and ATP regenerating enzyme. According to the method for regenerating ATP of the present invention, it has the following beneficial effects: (1) adopt three kinds of ATP regenerating enzymes of novel Ppk, Adk and Pap, the ATP consumed in the enzymatic reaction can be recycled and regenerated, greatly reducing the amount of ATP used, regenerated The process is simple and efficient, the reaction is easy to control, and the stability is high. (2) The substrate polyphosphoric acid or its salt to be added to the reaction is cheap and less polluting. (3) The enzymatic reaction does not need to add expensive ATP to start the reaction, add ADP or cheap AMP. (4) ATP regenerating enzyme recovery system was established, which is suitable for large-scale industrial production.

The invention discloses a method for preparing glutathione by means of an enzymic method. The method comprises the following steps that 1, Gsh F is utilized for generating glutathione in a reaction kettle; 2, regenerating enzyme is added into the reaction kettle, and ADP and / or AMP generated through a reaction is regenerated to be ATP; 3, the Gsh F and the regenerating enzyme which are fixed are separated in the reaction kettle or a filter device is used for separating dissociative Gsh F and the regenerating enzyme; 4, the product GSH and ATP, ADP and AMP in a reaction solution are separated. According to the preparation method, Gsh F enzyme is used for replacing Gsh I enzyme and Gsh II enzyme, a two-step reaction is changed into the one-step reaction, time is saved, and the feedback inhibition of the GSH for the two-step reaction is lowered; an ATP regenerating system for GSH production is established, the ATP consumption is lowered, and the cost is lowered; impurities such as pigment introduced by ATP regenerated by the yeast are avoided, and purification is easy; the enzyme recycling system is built, and large-scale production can be achieved.

The invention discloses a method for preparing glutathione with an enzymatic method. The method includes the following steps that (1), the glutathione is generated in a reactor through GshF enzymes, ATP regenerated enzymes and AK enzymes; (2), the fixed GshF enzymes, ATP regenerated enzymes and AK enzymes are separated in the reactor, or unbonded GshF enzymes are separated through filtering equipment; (3), filtrate obtained in the step (2) is separated, coarse GSH and a small amount of ATP, ADP and AMP are obtained; (4), the GSH obtained in the step (3) is concentrated, crystallized and dried,and finished GSH is prepared. The method includes the following steps that adenosine is used for replacing the ATP or the AMP, and a large amount of cost is reduced for industrial production; reaction conditions of GSH production are optimized, and the reaction rate is high; a stable enzyme recovery system is established, and energy saving and environmental protection are achieved; byproducts canbe directly used for a circulating reaction, or collected in a centralized mode to be used for producing the ATP, and the operation is simple.

The invention discloses a production method for enzymatic reaction by utilizing adenosine to replace ATP and application thereof. The method comprises the following steps that (1) in an enzymatic reaction system, ATP regeneration enzyme, AK enzyme and the adenosine are added in proportion for enzymatic reaction; (2) immobilized ATP regeneration enzyme and AK enzyme are directly separated in a reaction tank, and free ATP regeneration enzyme and AK enzyme are subjected to ultrafiltration membrane separation in a filter; (3) percolate in step (2) is separated and purified to obtain a product. Theproduction method for enzymatic reaction by utilizing the adenosine to replace the ATP and the application thereof have the advantages that a large amount of cost is saved for industrial production by utilizing the adenosine to replace the ATP or AMP; a stable enzyme recovery system is established, and the energy conservation and environment protection are achieved; a small quantity of generatedby-products ATP, ADP and AMP are directly used for cyclic reaction, or used for producing the ATP, or purified intensively through filtration, ion exchange and other methods, and the operation is simple.

The invention discloses a method of enzymatic co-production. The method includes the following steps: adding two or more of reaction substrates, salt ions and adenosine in proportion in and enzymaticreaction system; and adding an ATP regeneration enzyme, an AK enzyme and two or more of production enzymes to the system for reaction. The method of enzymatic co-production has the following advantages: 1) two or more of the reaction substrates and corresponding production enzymes are added into the reaction system requiring ATP to generate a plurality of products with economic value, which simplifies a reaction process and has higher industrial value; 2) the co-production saves the ATP regeneration enzyme and AK enzyme and reaction time and reduces water, electricity and labor costs; and 3) the adenosine is used to replace conventional ATP or AMP, which saves a plurality of raw material costs for industrial production, and the adenosine has the advantages of low price and wide range of sources.

The present invention discloses a method for synthesizing glutathione by enzymatic catalysis, comprising the following steps: S1, mixing Upsilon-glutamylcysteine synthetase liquid and glutathione synthetase liquid to obtain mixed liquid A, mixing the mixed liquid A with acetate kinase to obtain mixed liquid B; S2, adding an immobilized carrier in the mixed liquid B, stirring for immobilizing, and filtering to obtain an immobilized enzyme; S3, blending glutamic acid, L-cysteine, glycine, magnesium sulfate, ATP (adenosine triphosphate) and acetyl phosphate into a reaction liquid, adding the immobilized enzyme, and stirring for reacting; S4, after reacting, filtering the reaction liquid, and extracting and refining filtrate to obtain the glutathione. The method of the invention has high synthetic efficiency, the production content of the reaction liquid is higher than 10 g / L, and substrate conversion rate reaches higher than 90%; by introducing acetate kinase to construct an ATP regeneration and coupling system, ATP charge is greatly reduced and production cost is significantly reduced; the immobilized enzyme is reusable and highly stable.

The invention relates to a method for regenerating ATP(adenosine triphosphate). A heterogeneous source is used for expressing polyphosphate kinase, and under catalyzing of polyphosphate kinase, polyphosphates with the low polymerization degree are used as phosphoric acid donors for regenerating ATP. Compared with an existing ATP regeneration method, adopted polyphosphate kinase is a normal-temperature enzyme, can be combined with multiple enzymes for promoting reaction coupling, and adopts a tripolyphosphate, a tetrapolyphosphate and a hexametaphosphate as the phosphoric acid donors, these polyphosphates are low in polymerization degree, common, easy to obtain and low in price, on the basis of simplifying the ATP regeneration process, feasibility of ATP regeneration is improved, and the ATP regeneration cost is reduced; activity of a PPK enzyme is not inhibited by polyphosphate concentration, and possibility is provided for industrialized large-scale production.

The invention provides a chassis system for ATP (Adenosine Triphosphate) regeneration and application. The chassis system for ATP regeneration comprises the following five enzymes: alpha-glucan phosphorylase, glucophosphomutase, phosphoglucose Isomerase, phosphoketolase and acetokinase. In addition, the chassis system can further comprise the following seven enzymes: transaldolase, transketolase,ribose-5-phosphate isomerase, ribulose-3 epimerase, triosephosphate isomerase, fructose-bisphosphate aldolase and fructose-1,6-bisphosphatase. The chassis system can be coupled to an enzymic catalyticreaction needing ATP, utilizes starch or maltodextrin as energy, is not added with any coenzyme, can efficiently regenerate the ATP at low cost through a one-pot reaction and is an economical, highlyefficient, stable and sustainable ATP regeneration system.

The invention provides a method for synthesizing galacto-N-biose, which belongs to the technical field of bioengineering and oligosaccharide synthesis. According to the method, a multi-enzyme catalytic system which is good in biological safety and wide in application is utilized, and an ATP regeneration circulating system is introduced into a multi-enzyme reaction system, so that the galacto-N-biose synthesis and the substrate utilization rate are improved. The invention provides the novel synthetic route for galacto-N-biose, lays a foundation for large-scale industrial production of galacto-N-biose, and has important economic value and social benefit. Meanwhile, the synthesis method is efficient, mild, simple, convenient, easy to implement, low in cost and suitable for industrial production, and has good practical application value.

The invention relates to Issatchenkia orientalis and a method for producing citicoline by whole cell conversion of Issatchenkia orientalis, belonging to the technical field of biological pharmacy. In the method provided by the invention, the whole cells of Issatchenkia orientalis Z1, namely CCTCC (China Center for Type Culture Collection) NO: M2011272 are utilized to prepare citicoline, choline phosphate and 5'-cytidylic acid are used as substrates, glucose is used as an energy donor, and the ATP (adenosine triphosphate) regeneration efficiency is improved by adding inorganic ions; glucose is used as the energy donor, so that the energy requirement of the strain is provided and ATP is provided for a citicoline synthesis enzyme system; one or more of potassium ion, magnesium ion and manganese ion are added to change the metabolism flow direction and improve the ATP regeneration rate, so that the ATP regeneration rate is matched with the rate of the citicoline enzyme synthesis system and the high-efficiency preparation of citicoline is achieved; and the whole cells of Issatchenkia orientalis are used, and toluene is added to an aqueous solution during the preparation process so as to improve the cell permeability, so that the rate of the citicoline enzyme synthesis system is improved.

The invention discloses a catalytic system coupled with an ATP regeneration system and application of the catalytic system in glutathione production. The application includes: building heterogenous expression / genome integrated Escherichia coli containing glutathione synthesis difunctional enzyme and ATP regeneration enzyme, performing glutathione synthesis reaction condition and catalytic system optimization, and using the mixed culture of the two kinds of Escherichia coli and strain proportion and catalytic reaction process optimization to achieve glutathione production, and raw material, equipment and synthesize reaction costs are saved to the maximum extent.

The invention relates to a method for producing glutathione, which comprises the following steps: doing large scale culture and separating purify technology to two types of expressing recombining gene's gene project bacillus coli with glutathione enzyme A and enzyme B to obtain enzyme A and enzyme B and doing solid to them by suit materials (1); mixing the two solid enzymes with the ratio EA: EB=1:10-10:1 into the GSH synthesis reactor, which is used in the synthesis of glutathione (2); forming the total producing system into a large fluid cycle type reaction system because GSH synthesis reactor and ATP re-generating reactor are fluid cycle(3); the aminothiopropionic acid availability ratio is above 40%; the glutathione recovery ratio s above 50%; the main synthesis reacting stage can hold the density of glutathione above 0.8 g / L.

The invention relates to a preparation method of sedoheptulose and aldose in an immobilized enzyme cascade reaction, and specifically discloses a method for preparing sedoheptulose and aldose in the immobilized enzyme cascade reaction. The method comprises the following steps: taking L-erythrulose and 1,3-dioxyacetone as starting materials, and preparing D-sedoheptulose-7-phosphoric acid by usinga one-pot method of immobilized kinase EK, immobilized isomerase TRI, erythrulose phosphoric acid isomerase RPI, immobilized aldolase FSA and immobilized ATP regenerated enzyme PPK; and further generating sedoheptulose and aldose. The method is high in preparation efficiency and cheap in raw materials, and is applicable to large-scale industrial production, so that the preparation cost is greatlyreduced.

The invention relates to a method for measuring 1,5-anhydroglucitol by oxidase, which can be widely applied in the field of medical and biochemical technologies. The method comprises the following steps of: converting glucose (Glu) in a sample into fructose-6-phosphate (Fru-6-P) by glucokinase (GK) and phosphoglucose isomerase (PGI), and establishing an ATP regeneration system by pyruvate kanise (PK) and phosphoenolpyruvate (PEP) in the system so as to ensure that Glu is converted into Fru-6-P. In the sample, 1,5-AG reacts in the presence of pyranose oxidase (PROD) to produce 1,5-fructosan (1,5-AF) and H2O2, and then H2O2 is colored and quantified via a Trinder's system..

The invention relates to a method for regenerating ATP using a rationally designed enzyme. Through rational design after sequence alignment, polyphosphate kinase from sinorhizobium meliloti is mutated and heterologously expressed; and under the catalysis of the polyphosphate kinase, the regeneration of ATP is realized by adopting polyphosphate with low polymerization degree as a phosphoric acid donor. Compared with existing ATP regeneration method, the polyphosphate kinase used in the invention is a normal-temperature enzyme which can react for coupling with multiple enzymes; the enzyme can take tetra-polyphosphate as a phosphoric acid donor; compared with phosphoric acid with high polymerization rate, the tetra-polyphosphoric acid is more common and easily available; and the feasibility of ATP regeneration is increased on the basis of simplifying the ATP regeneration process, and the cost of ATP regeneration is reduced. The activity of PPK enzyme is not inhibited by the polyphosphate concentration, and industrial large-scale production is possible.

The utility model discloses a surface cleanliness detection reagent used in health supervision, which is based on the combination of ATP biological fluorescence method and enzymatic cycling amplification technology to realize rapid detection and field assessment of health surface cleanliness. The utility model comprises ATP releaser and a fluorescent inducer with ATP regeneration enzyme; wherein, the ATP releaser can release microorganisms, somatic cells and ATP in product residues; the fluorescent inducer can react with the ATP to produce quantitative fluorescence, and then combined with the ATP regeneration enzyme to amplify the produced fluorescent signals further. The utility model, which has the advantages of convenient use of reagent, simple operation, fast response, high sensitivity and low detection cost, is widely used in health monitoring and health field supervision in food industry, cosmetic industry, medical industry and other industries.

A method of producing a substance includes synthesizing a molecule at least by mixing substrates, a synthase, adenosine triphosphate (ATP), a polyphosphate kinase 2, and a polyphosphoric acid mixture. The polyphosphoric acid mixture includes 50% or more of polyphosphoric acid with a degree of polymerization of not less than 15. Adenosine diphosphate (ADP) is generated from the ATP during the synthesis. The synthesis is coupled with an ATP regeneration reaction in which the ATP is regenerated by the polyphosphate kinase 2 from the ADP and the polyphosphoric acid.

The invention discloses a method for preparing a theanine by an interspecific coupling ATP regenerative technology comprising the following steps: preparing gamma-glutamyltransferase immobilized cells by using bacillus licheniformis, preparing 1,6-diphosphate conversion culture solution by using bacillus stearothermophilus; preparing the theanine conversion solution by mixing the gamma-glutamyltransferase immobilized cells, 1,6-diphosphate conversion culture solution and substrates which are glutamine (glutamic acid) and ethylamine; obtaining the theanine by extracting and refining the theanine conversion liquid. The technical scheme provided by the invention is a fine new technical scheme for theanine preparation by coupling the ATP regenerative technology and an enzyme conversion technology, a technology capable of promoting the ATP consumed enzyme reaction smooth, also a technology suitable for the theanine production by the microorganism conversion method and has a wide application prospect in the food, medical treatment and the enzyme engineering field.

The invention discloses a polyphosphate kinase mutant, an engineering bacterium and application of the polyphosphate kinase mutant and the engineering bacterium. The polyphosphate kinase mutant is obtained by performing single mutation or multiple mutation on the 79th site, the 106th site, the 108th site, the 111th site or the 285th site of an amino acid sequence shown in SEQ ID NO: 2. The invention provides a plurality of polyphosphokinase mutants derived from Cytophaga hutchinsonii, the specific enzyme activity of the mutants is increased by 2.7-17.9 times compared with that of female parent polyphosphokinase, an ATP regeneration system formed by the mutants can reduce the consumption of ATP in ATP-dependent biocatalytic synthesis reaction by 70% or more, and the mutant has a wide industrial application prospect.

The present invention discloses a production method of enzymatic reaction using adenosine instead of ATP. The method comprises the following steps: (1) adding ATP regeneration enzyme, AK enzyme and adenosine in proportion to carry out an enzymatic reaction in an enzymatic reaction system; (2) separating the ATP regeneration enzyme and AK enzyme by either directly separating ATP regeneration enzyme and AK enzyme immobilized in a reaction tank or separating free ATP regeneration enzyme and AK enzyme by an ultrafiltration membrane in a filter; and (3) separating and purifying the filtrate of step (2) to obtain a product. The disclosed method provides: greatly reduced industrial production costs; faster reaction rate; stable enzyme recovery system that is energy efficient and environmentally friendly; and capability of reusing the byproducts or collecting them for the production of ATP.

The invention discloses an efficient catalytic synthesis method of PAPS based on construction of an ATP regeneration system, and belongs to the technical field of bioengineering. PAPS is artificially constructed through microbial recombinant expression to synthesize a bifunctional enzyme, and high-efficiency production of PAPS is realized. On the basis, an ATP regeneration system of polyphosphokinase from corynebacterium glutamicum and mycobacterium tuberculosis is coupled, two by-products including pyrophosphoric acid and ADP can be recovered at the same time, equivalent conversion of a substrate and a product is achieved, PAPS generated in a catalytic system has high purity, and supply of sulfonic acid groups in most sulfonic acid transfer reactions can be achieved.

The present invention discloses an enzymatic method for preparing glutathione (GSH), comprising the following steps: (1) producing glutathione in a reactor using GshF enzyme, ATP regeneration enzyme and AK enzyme; (2) separating immobilized GshF enzyme, ATP regeneration enzyme and AK enzyme in the reactor, or separating free GshF enzyme using a filtration device; (3) separating the filtrate obtained in step (2) to obtain a crude product GSH and a small amount of ATP, ADP and AMP; and (4) subjecting the GSH obtained in step (3) to concentration, crystallization, and drying to prepare a finished GSH. The disclosed method provides: greatly reduced industrial production costs; faster reaction rate; stable enzyme recovery system that is energy efficient and environmentally friendly; and capability of reusing the byproducts or collecting them for the production of ATP.

The invention discloses a method for synthesizing D-psicose by combining phosphorylation and an ATP (adenosine triphosphate) regeneration system, which comprises the following steps: by taking D-glucose as a raw material, sequentially carrying out recombinant escherichia coli construction, wet cell preparation, whole-cell catalytic reaction and dephosphorylation to prepare the D-psicose. Wherein the construction process of the recombinant escherichia coli comprises the step of introducing an ATP regeneration system gene. According to the method, the equilibrium conversion rate of synthesizing the D-psicose from the D-glucose is increased, and the addition of exogenous ATP (adenosine triphosphate) is reduced. The method has the characteristics of simple preparation process, high product conversion rate, greenness and economy, and can be used for industrial production.

The invention relates to a method for measuring 1,5-anhydroglucitol by dehydrogenase, which can be widely applied in the field of medical and biochemical technologies. The method comprises the following steps of: converting 1,5-anhydroglucitol (1,5-AG) to be measured and a main interferent, i.e., glucose (Glu), into 1,5-anhydroglucitol-6-phosphate (1,5-AG-6-P) and glucose-6-phosphate (Glu-6-P) by ATP dependent hexohinase (ATP-HK), respectively,and establishing an ATP regeneration system by piruvate kinase (PK) and phosphoenolpyruvate (PEP) in the system so as to ensure that 1,5-AG and Glu is converted into 1,5-AG-6-P and Glu-6-P, respectively. The converted 1,5-A-G-P is specifically dehydrogenated and oxidized by 1,5-anhydroglucitol-6-phosphate dehydrogenase (1,5-AG-6-PDH), and 2-(4-iodophenyl)-5-(2,4-dinitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazole is reduced to a colored substance and then quantified by a full automatic biochemical analyzer by colorimetry.

A method for cell-free protein synthesis is characterized in that pH is controlled by using an enzyme. For example, by using an amino acid decarboxylase, the pH is controlled according to removal of hydrogen ions that are produced during regeneration of ATP. The method for cell-free protein synthesis of the present invention has an advantage that not only the expression amount of protein is enhanced but also the expressed protein can be directly used for activity analysis without undergoing any separation or purification.

The present invention provides for a nutritional composition in a liquid drink form, dry powder form, gel form, or carrier form intended to optimize human physiological performance during intense exercise and subsequently return the body to its healthy, pre-exercise state. The composition comprises a carbohydrate and a protein, and preferably a ratio of the carbohydrate to the protein is greater than about 4.3 parts by weight carbohydrate to 1 part by weight protein. Preferably, the liquid nutritional composition is comprised of: (1) carbohydrate in the range of 3.45-10% by weight of the liquid nutritional composition to allow immediate and long-term energy production by muscle cells and neurons, insulin modulation, and muscle glycogen sparing, the carbohydrate being in the form of monosaccharides or polysaccharides or a combination thereof, and the saccharides preferably comprising 40-95% dextrose by weight of the final volume of saccharides, 5-45% fructose by weight of the final volume of saccharides, and 1-25% glucose polymer by weight of the final volume of saccharides; (2) a protein source that contains all 20 “standard” amino acids, in the range of 0.1-0.8% by weight of the liquid nutritional composition, whose inherent structure plus further chemical modification results in amino acid absorption into the bloodstream rapid enough for initiating and enhancing protein synthesis and subsequent muscle cell repair during the exercise and post-exercise periods; (3) Branched-Chain Amino Acids (BCAA), leucine, isoleucine, valine, for use by muscle cells for energy thus sparing glycogen stores, for increasing protein synthesis by stimulating insulin and growth hormone release, and for reducing protein degradation; (4) precursors of the signaling molecule nitric oxide, important for increasing blood flow and subsequent nutrient delivery (including those in this composition) to tissue during intense exercise; (5) precursors of creatine, a molecule important for ATP regeneration in muscle and nerve cells and for muscle cell volumizing effects; (6) a mixture of hydrophilic and hydrophobic antioxidants: retinol / Beta-carotene (Vitamin A), ascorbic acid (Vitamin C), and di-alpha-tocopheryl acetate (Vitamin E); ubiquinone (coenzyme Q10 [5-100 mg / 500 ml], which in addition to its antioxidant properties is also important for ATP generation and Vitamin E regeneration; a mixture of 10-100 mg epigallocatechin gallate (EGCG) and 25-150 mg polyphenols from green tea extract; 40-250 mg ellagic acid and punicalagin from pomegranate or raspberry extract, at ratios comparable to that found in the whole fruit, which have been shown to have protective effects on blood vessels as well as inhibiting stimulation of the pro-inflammatory protein interleukin-1b (IL-b) which causes degradation and damage to joints (These six antioxidants prevent and eradicate reactive oxygen species and free radicals generated during strenuous exercise, thus preserving muscle integrity.); and (7) the ions Na+, Cl−, K+, Ca+2, Mg+2, and Zn+2, which are important for the muscle fiber contraction and relaxation cycle, for neuronal signal propagation, for repletion of such ions lost in excessive perspiration during exercise, for water absorption in the gastrointestinal tract, and for catalytic action by energy producing and antioxidant enzymes.

The amino acid sequence of the polyphosphate kinase mutant is SEQ ID NO: 3, the enzyme activity of the mutant is improved by at least three times compared with that of wild enzyme, and the polyphosphate kinase mutant can be used for improving the ATP regeneration capacity of microorganisms.

The present invention provides a kind of synthetic milk- N A method for disaccharides belongs to the technical field of bioengineering and oligosaccharide synthesis. The present invention utilizes the multi-enzyme catalytic system with good biological safety and wide application, and improves milk- N ‑Disaccharide synthesis and substrate utilization. The present invention provides a new milk- N ‑disaccharide synthesis pathway for milk‑ N The large-scale industrial production of disaccharides has laid the foundation and has important economic value and social benefits. Simultaneously, the synthesis method of the present invention is efficient, mild, simple and easy to implement, low in cost, suitable for industrialized production, and has good practical application value.

A method of providing electrons to an electron transport chain capable of generating a proton gradient to regenerate ATP from ADP using an electrochemical cell, in particular a membrane bioreactor. Such electron transport chains may be part of a synthetic membrane, or included within the synthetic membrane, or may be prepared by appropriately disrupting living cells. The electrons provided by the electrochemical cells are delivered to the electron transport system through suitable electron carriers, such as NADH2, FMNH2, FADH2, reducing ubiquinone, thiol or other electron carriers or biologically reducing equivalents compatible with electron transport chain components that enable ATP regeneration.