42 results about "Purine nucleoside phosphorylase" patented technology

The invention relates to the field of an external diagnostic reagent, and particularly relates to a high-sensitivity kit for detecting 5'-nucleotidase. The kit for detecting 5'-nucleotidase comprises a 5'-nucleotidase R1 reagent, a 5'-nucleotidase R2 reagent and a 5'-nucleotidase calibration product, wherein the 5'-nucleotidase R1 reagent comprises a buffer solution, a stabilizing agent, a preservative agent, an enzyme activator, 4-AAP(4-aminoantipyrine), 18-crown ether-6 and UAO (Uricase), PNP (Purine Nucleoside Phosphorylase), XOD (Xanthine Oxidase) and POD (Peroxidase) reaction enzyme systems; the 5'-nucleotidase R2 reagent comprises a buffer solution, a stabilizing agent, a preservative agent, PO3- and a Trinder developing system; and the 5'-nucleotidase calibration product comprises a buffer solution, a stabilizing agent, a preservative agent and 5'-nucleotidase. The detection kit provided by the invention is enhanced in detection sensitivity by 40% compared with a 'PNP-XOD-POD' enzyme system, and has good application prospects.

Purine N9-β-D-nucleosides containing 3-amino-3-deoxy-β-D-ribofaranose, 3-amino-2,3-dideoxy-β-D-ribofuranose, and 2-amino-2-deoxy-β-D-ribofuranose as sugar moieties are synthesized by biocatalytic transglycosylation of purine bases and the respective 3′-amino-3′-deoxyuridine, 3′-amino-3′-deoxythymidine and 2′-amino-2′-deoxyuridine as donors of the carbohydrate moiety, and the cells of Escherichia coli as a biocatalyst or glutaraldehyde (GA) treated cells of Escherichia coli as a biocatalyst or a mixture of thymidine (uridine) phosphorylase and purine nucleoside phosphorylase.

The invention discloses a uridine phosphatase mutant and a preparation method thereof. The amino acid sequence of the mutant is SEQ ID NO: 3, ribose-1-phosphoric acid can be effectively catalyzed to react with nicotinamide to generate nicotinamide ribose; the nicotinamide ribose can form a three-enzyme system together with purine nucleoside phosphorylase and nicotinamide ribose kinase to jointly catalyze a reaction of raw materials guanosine, phosphate, nicotinamide and ATP to synthesize the beta-nicotinamide mononucleotide by a one-pot method, and the product has good industrial development and application prospects.

The present invention provides engineered purine nucleoside phosphorylase (PNP) enzymes, polypeptides having PNP activity, and polynucleotides encoding these enzymes, as well as vectors and host cellscomprising these polynucleotides and polypeptides. Methods for producing PNP enzymes are also provided. The present invention further provides compositions comprising the PNP enzymes and methods of using the engineered PNP enzymes. The present invention finds particular use in the production of pharmaceutical compounds.

The invention relates to an adenosine deaminase detection kit. A vessel is contained in the detection kit and filled with liquid for detection, the liquid for detection is prepared from 50-100 mmol / L of glycine buffer solution, 50-80 mmol / L of sodium benzoate, 12 / 18 u / L of purine nucleoside phosphorylase, 30-40 u / L of xanthine oxidase, 200-300 u / L of peroxidase, 30-100 g / L of glyceraldehyde, 0.5 g / L of sodium azide, 10-200 mmol / L of disodium hydrogen phosphate, 100-300 mmol / L of KCl, 40-100 mmol / L of mannitol, 0.1-0.3 ml / L of preservative, 100-150 mmol / L of adenosine, 2-4 mmol / L of 4-aminoantipyrine, 10-15 mmol / L of color developing agent, 20-40 mmol / L of adenine nucleoside and 3-5 g / L of bovine serum albumin. The detection kit has the advantages of being high in determination precision, high in antijamming capacity, suitable for automated rapid determination and capable of creating favorable conditions for routine development of ADA detection application in clinic.

The invention provides an assay kit for adenosine deaminase. The assay kit comprises a reagent R1 and a reagent R2. The reagent R1 comprises TRIS, HCL, xanthine oxidase, purine nucleoside phosphorylase, 4-aminoantipyrine, peroxidase and sodium aspartate; and the reagent R2 comprises TRIS, HCL, adenosine, N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3-methylaniline sodium slat and glycerol. The assay kit ofthe invention belongs to the technical field of biological detection, is simple in reagent composition, good in stability, rapid in reaction and low in cost, and can accurately detect adenosine deaminase; and the detection results of the assay kit have no obvious difference from the detection results of listed products.

The invention discloses a 5'-ribonucleotide hydrolytic enzyme detection kit. A kit body comprises a glycine buffer solution, a color developing agent prepared from 3- (m-tolidino) dipropane sulfonic acid disodium salt, purine nucleoside phosphorylase, xanthine oxidase, peroxidase, 5'-hypoxanthine nucleotide and 4-aminoantipyrine solution. A sample is mixed with a reagent according to a certain volume ratio for a series of reaction, reactants are placed under a semi- / full-automatic biochemical analyzer, and the change speed of light absorptivity in the position of a main wavelength of 550nm is detected so as to measure and calculate the concentration of the 5'-ribonucleotide hydrolytic enzyme. The detection kit has the advantages of accuracy, stability and convenience.

The invention discloses a method for preparing ribavirin through genetically engineered bacteria and belongs to the technical field of genetic engineering. Particularly, the method comprises the stepsthat a pun gene segment with a nucleotide sequence shown in SEQ ID NO:1 is synthesized; a recombinant plasmid pET20b-BA-pun is established, and then the genetically engineered bacteria BL21 / pET20b-BA-pun are established and induced so as to obtain a key enzyme, namely purine nucleoside phosphorylase, for synthesizing the ribavirin. The genetically engineered bacteria are used for synthesizing theribavirin, the conversion rate is high, the cost is low, and the method is environmentally friendly and has important significance in industrial production.

An adenosine deaminase assay kit consists of a reagent R1 and a reagent R2 which are independent from each other, wherein the reagent R1 comprises the following components: a buffer solution, a preservative, a protective agent, an anti-interference agent, a surfactant, a reaction enhancer, peroxidase, purine nucleoside phosphorylase, xanthine oxidase, 4-aminoantipyrine and water; and the reagent R2 comprises the following components: a buffer solution, a protective agent, a reaction enhancer, a preservative, adenosine, a chromogen substance and water. The adenosine deaminase assay kit is highin analysis sensitivity, good in repeatability and high in anti-interference capability, and the linearity can reach 300 U / L or above.

The invention discloses a liquid stable 5 '-nucleotidase calibrator. The calibrator comprises the following components: a buffer solution, a stabilizer, a preservative and 5'-nucleotidase. The invention also discloses a 5 '-nucleotidase detection kit. The kit comprises a 5'-nucleotidase R1 reagent and a 5 '-nucleotidase R2 reagent. The 5 '-nucleotidase R1 reagent is an enzyme reaction system and consists of a first buffer solution, a stabilizer, a first preservative, 4-aminoantipyrine, an enzyme activator, purine nucleoside phosphorylase (PNP), xanthine oxidase (XOD) and peroxidase (POD); the 5 '-nucleotidase R2 reagent is a chromogenic system and is composed of a second buffer solution, a second preservative, inosine-5'-disodium phosphate, beta-sodium glycerophosphate and a Trinder chromogenic substrate. The invention also discloses an application of the 5 '-nucleotidase detection kit in determination of 5'-nucleotidase.

The invention relates to a method for determining the activity of 5'-nucleotidase, further to a 5'-nucleotidase detection kit, and belongs to the technical field of medical test determination. According to the method, 5-inosine monophosphate used as a substrate is converted and dehydrogenated by using purine nucleoside phosphorylase and xanthine dehydrogenase as tool enzymes, oxidized coenzyme isreduced to form a reduced coenzyme, and the enzyme reaction rate is calculated by detecting the generation amount of the reduced coenzyme at a wavelength of 340 nm. Compared to the method in the priorart, the method of the present invention has advantages of less reaction steps, less kinds of raw materials, strong alkaline-phosphatase resistance, stable reagent performance, wide linear range, high sensitivity and good accuracy.

A method for producing cladribine (2-chloro-2′-deoxyadenosine) comprising the steps of:a) reaction of 2-deoxyuridine with 2-chloroadenine, in the presence of uridine phosphorylase (UPase) and purine nucleoside phosphorylase (PNPase) in an aqueous reaction medium possibly containing up to 40% v / v of an aprotic dipolar solvent, to obtain cladribine dissolved in said reaction medium;b) isolation of the cladribine by precipitation by means of concentration and alkalinisation of the reaction medium up to pH 11.5-12.5.

The invention discloses a detection reagent kit for detecting human ADA. A reagent kit body contains a glycine buffer solution, a color-developing agent prepared with 3-methyl-N,N-dipropyl sodium sulfonate aniline, purine nucleoside phosphorylase, xanthine oxidase, peroxidase, adenosine and a 4-aminoantipyrine solution. A sample and a reagent are mixed in a certain volume proportion and subjected to a series of reactions, then reactants are placed under a semi-automatic / full-automatic biochemical analyzer, and the change rate of the absorbency at a position of 550nm dominant wavelength is detected, so that the concentration of the ADA is measured and calculated. The reagent kit has the advantages of being accurate, stable and convenient.

The invention belongs to the technical field of medical and biochemistry, and particularly relates to an adenosine deaminase measuring kit. The kit is composed of a reagent R1 and a reagent R2, wherein the reagent R1 comprises biobuffer, preservative, peroxidase, 4-aminoantipyrine, purine nucleoside phosphorylase, xanthine oxidase and water, and the reagent R2 comprises biobuffer, preservative, adenosine, N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3-methylaniline and water. The adenosine deaminase measuring kit conducts a test according to a continuous monitoring method, the main wavelength is 550 nm, and the kit has the advantages of being quick, sensitive, high in accuracy, specificity and stability and the like.

The invention provides a staphylococcus sp. LZ16 strain, which has a collection number of CGMCC (China General Microbiological Culture Collection Center) No.6121. The staphylococcus sp. contains Purine nucleoside phosphorylase (PNP) protein and has an inhibiting effect on plant pathogenic bacteria. The amino acid sequence of the staphylococcus sp. is disclosed in SEQ ID (Identity) No.3, and the nucleotide sequence of a gene for encoding the antibacterial protein is disclosed in SEQ ID No.2. The staphylococcus sp. LZ16 strain provided by the invention has an obvious inhibiting effect on the pyricularia oryzae. Antibacterial agent or biopesticide prepared from the strain LZ16 disclosed by the invention can generate obvious acceleration action on preventing and curing the pyricularia oryzae,the staphylococcus sp. LZ16 strain can be popularized and used in agricultural production, and the production performance of paddy is effectively improved.

The invention belongs to the technical field of medicine and biochemistry and particularly relates to an adenosine deaminase detection kit. The kit comprises a reagent R1 and a reagent R2; the reagentR1 comprises purine nucleoside phosphorylase, xanthine oxidase, peroxidase, a preservative, 4-amino antipyrine and a buffer solution; the reagent R2 comprises adenosine, a color developing agent, a preservative and a buffer solution. The adenosine deaminase detection kit adopts a continuous monitoring method for detection, and the main wavelength is 550 nm; the detection kit has the advantages ofhigh sensitivity and accuracy and the like.

The invention discloses a kit for determining 5'-ribonuclease. The kit is composed of a reagent R1 and a reagent R2 which are independent of each other. The reagent R1 is prepared from the following components: a buffer solution, a preservative, a protective agent, an anti-interference agent, a surfactant, a reaction enhancer, peroxidase, purine nucleoside phosphorylase, xanthine oxidase, a chromogen substance and water; and the reagent R2 is prepared from the following components: a buffer solution, a protective agent, a reaction enhancer, a preservative, disodium inosinate, 4-aminoantipyrineand water. The kit is high in analysis sensitivity, good in repeatability and high in anti-interference capability, the linearity can reach up to 400U / L, the detection of abnormal specimen results conforms to the actual concentration, and the accuracy of normal specimen test results is not affected.

The invention discloses the testing method of adenosine deaminase. The testing reagent of adenosine deaminase is made up of the following two parts: Reagent 1 includes disodium hydrogen phosphate 8.5-12 mmol / L, aminoantipyrine 1.5-2.2 mmol / L, purine nucleoside phosphorylase 2500-2900 U / L, xanthine oxidase 500-700 U / L, peroxidase 510-710 U / L, ascorbinase 1800-2800 U / L, certain stability reagent and certain assistant factors. Reagent 2 includes adenosine 10-14 mmol / L and EHSPT 1.5-2.5 mmol / L. The testing reagent formula of adenosine dehydrogenase in this invention and the confirmation of the testing parameters and referential range are good for the consistency of the clinical testing results. it will bring a lot of benefits to the clinical diagnosis. The sensitivity of this method is very high and it is superior to the similar products sold on the market.

The invention relates to a method for synthesizing vidarabine by an enzymic method, and provides a method for preparing vidarabine by a two-step method. According to the method, different functions of uridine phosphorylase and purine nucleoside phosphorylase are utilized fully, so that reaction is carried in one direction; reverse reaction is eliminated; and conversion rate of the vidarabine is increased remarkably.

The invention discloses a method for measuring relative content of hypoxanthine riboside and adenosine in samples to be tested. The method includes converting hypoxanthine riboside in an adenosine sample into purple red quinone derivatives by an enzyme-coupled system composed of PNP (purine nucleoside phosphorylase), XTO (xanthine oxidase) and POD (peroxidase), and measuring solution absorbance value A1 under specific wavelength; measuring absorbance value A2 after adding buffer solution containing adenosine deaminase and keeping reaction for some time, and calculating relative content of hypoxanthine riboside and adenosine in adenosine substrate by the values A1 and A2. The method is green and environment friendly, relative content of hypoxanthine riboside and adenosine can be calculated without reference of calibrators, and detection can be realized by a common visible spectrophotometer. The method is high in flexibility, good in linearity and precision, accurate in results and convenient and quick to operate, and can be used for various types of analyzers to meet requirements of testing the samples in large scale.

The invention discloses an apolipoprotein E test kit, comprising a reagent R1 and a reagent R2. The reagent R1 comprises a glycine buffer solution, polyoxyethylene, sodium ethylene diamine tetracetate, purine nucleoside phosphorylase, dodecyl phenol, xanthine oxidase, sodium benzoate, glyceraldehyde, potassium dihydrogen phosphate. The reagent R2 comprises a glycine buffer, a latex particle of apolipoprotein E monoclonal antibody, nonidet p40, potassium chloride, calcium acetate, adenosine, disodium hydrogen phosphate, sodium ethylene diamine tetracetate, hypoxanthine nucleotide. The apolipoprotein E test kit can accurately measure in vitro, and is effectively applied to clinical auxiliary diagnosis and screenings for diseases such as hyperlipemia, coronary heart disease, hepatopathy, acute hepatitis, biliary cirrhosis, etc., and has excellent detecting stability and detecting accuracy.

The present invention relates to a 2'-deoxy-2'-fluoro-beta-D-arabinofuranosyladenine production method, and discloses mutations at a certain specific loci of the amino acid sequences of purine nucleoside phosphorylase (PNP) and pyrimidine nucleoside phosphorylase (PyNP), and a series of obtained mutants, wherein the transglycosylation activities of the mutants are extremely significantly improved.

The present invention provides engineered purine nucleoside phosphorylase (PNP) enzymes, polypeptides having PNP activity, and polynucleotides encoding these enzymes, as well as vectors and host cells comprising these polynucleotides and polypeptides. Methods for producing PNP enzymes are also provided. The present invention further provides compositions comprising the PNP enzymes and methods of using the engineered PNP enzymes. The present invention finds particular use in the production of pharmaceutical compounds.

A glyoxal-guanosine-group compound is prepared either by reacting glyoxal-guanine with any one of ribose-1-phosphate and 2-deoxyribose-1-phosphate in the presence of purine nucleoside phosphorylase, or by reacting glyoxal-guanine with any one selected from the group consisting of uridine, 2′-deoxyuridine and thymidine, together with phosphate ion, in the presence of purine nucleoside phosphorylase and pyrimidine nucleoside phosphorylase. The glyoxal-guanosine-group compound is then decomposed by alkali, whereby a guanosine-group compound consisting of guanosine and 2′-deoxyguanosine is prepared.

The invention is related to phosphorus substituted purine nucleoside phosphorylase inhibitory compounds, compositions containing such compounds, and therapeutic methods that include the administration of such compounds, as well as to processes and intermediates useful for preparing such compounds.

The invention relates to a method for enzymatically synthesizing vidarabine. The invention provides a two-step method for preparing vidarabine adenosine. The method makes full use of the different effects of uridine phosphorylase and purine nucleoside phosphorylase to make the reaction proceed in one direction, eliminate the occurrence of reverse reaction, and significantly improve The conversion rate of vidarabine.

The invention relates to a method for determining the activity of adenosine deaminase, further to an adenosine deaminase detection kit, and belongs to the technical field of medical test determination. According to the method, adenosine used as a substrate is converted and dehydrogenated by using purine nucleoside phosphorylase and xanthine dehydrogenase as tool enzymes, oxidized coenzyme is reduced to form a reduced coenzyme, and the enzyme reaction rate is calculated by detecting the generation amount of the reduced coenzyme I at a wavelength of 340 nm. The method of the present invention has advantages of less reaction steps, less kinds of raw materials, good reagent stability, wide linear range, high sensitivity, good accuracy and convenient promotion.

The invention relates to primers, kit and method to detect transcriptional level of purine nucleotide phosphorylase PNP gene in macaque liver by RT-qPCR (real-time quantitative polymerase chain reaction), and belongs to the technical field of molecular biology. A template is made with cDNA synthesized by reverse transcription of total RNA extracted from fresh macaque liver tissue; a PCR primer combination is used to perform real-time fluorescent quantitative PCR amplification; Ct values of PNP gene fragments and reference gene GAPDH fragments, dissolution peaks, and amplification efficiency are attained; multiple-expressed delta delta C(t) value of the homogenize PNP gene is acquired by conventional treatment; relative expression value of the transcriptional level of PNP gene is acquired finally. An effective means to study PNP functionality of macaque and influence of relevant drugs upon PNP is provided; in addition, the primers, kit and method herein are applicable to real-time quantitative PCR detection and have the advantages of good operational simplicity, high repeatability, low detection cost, high sensitivity, high specificity and good convenience of popularization and application.

The invention discloses a uridine phosphatase mutant whose amino acid sequence is SEQ ID NO: 3, which can effectively catalyze the reaction of ribose-1-phosphate and nicotinamide to generate nicotinamide ribose, and can be combined with purine nucleoside phosphorylase and Nicotinamide ribokinase together constitutes a three-enzyme system and catalyzes the reaction of raw materials guanosine, phosphate, nicotinamide and ATP to synthesize β-nicotinamide mononucleotide in one pot, which has good industrial development and application prospects.