39 results about "Oxalate decarboxylase" patented technology

It is intended to provide a means of highly producing oxalate decarboxylase originating in a microorganism. A recombinant expression plasmid vector, which contains a-amylase promoter of a microorganism belonging to the strain Bacillus and an oxalate decarboxylase gene originating in a microorganism that is provided under the control of the promoter, is constructed. A host bacterium is transformed by this vector to give an oxalate decarboxylase-producing bacterium. A recombinant oxalate decarboxylase is produced by culturing the producing bacterium as described above and then harvesting the oxalate decarboxylase thus produced.

Oxalate decarboxylase crystals, including stabilized crystals, such as cross-linked crystals of oxalate decarboxylase, are disclosed. Methods to treat a disorder associated with elevated oxalate concentration using oxalate decarboxylase crystals are also disclosed. Additionally disclosed are methods of producing protein crystals.

The invention discloses glycosylated oxalate decarboxylase. The oxalate decarboxylase is derived from edible basidiomycete; the glycosylated oxalate decarboxylase has enzymatic activity in an environment that pH value is 1.5-7.5; and the activity is 10% greater than optimum activity in an environment that pH value is 1.5-2.0, which is 2U / mg greater than an activity unit. The invention also discloses a preparation method and an application of the glycosylated oxalate decarboxylase. The glycosylated oxalate decarboxylase provided by the invention can keep activity under a low-pH condition; and the glycosylated oxalate decarboxylase can be used for effectively preventing and treating kidney stone.

The invention discloses a recombinant plasmid for production of oxalate decarboxylase, a system and a method for expressing escherichia coli, and application. The recombinant plasmid for production ofoxalate decarboxylase comprises an oxalate decarboxylase gene, a molecular chaperone gene and a gene for regulating and controlling intracellular manganese ion concentration. The recombinant plasmidis guided into an escherichia coli expression bacterial strain and the escherichia coli with MntP gene deletion to respectively obtain oxalate decarboxylase with solubility expression and activity; the culture temperature and induction temperature of the recombinant bacterial strain are optimized; when the culture temperature is 37 DEG C, and the induction temperature is 25 DEG C, the expression amount of the oxalate decarboxylase is higher; by utilizing the two steps of organic solvent precipitation and ammonia sulfate precipitation to simply purify, the specific activity of the oxalate decarboxylase can reach 45.6U / mg, and the activity is higher than 95%. By adopting the technical scheme, the recombinant plasmid has the advantages that the production and purifying technologies are simple; the expression amount and specific activity of the oxalate decarboxylase are higher, the industrialized amplification is easy, the cost is low, and the recombinant plasmid is suitable for the industrialized production and application of the oxalate decarboxylase.

The invention provides a method for cross-linking and fixing oxalate decarboxylase by cross-linked oxalate decarboxylase aggregates (CLEAs), belonging to the technical field of biochemical engineering. The method comprises the following steps: induced expression of recombinant oxalate decarboxylase, separation and purification of oxalate decarboxylase, and preparation of CLEAs. The invention is characterized in that ethanol, acetone or ammonium sulfate are utilized to carry out precipitation separation on oxalate decarboxylase in a crude enzyme liquid; ethanol is utilized to prepare the oxalate decarboxylase aggregates; 0.01-0.15% glutaric dialdehyde is utilized to carry out cross-linking reaction for 0.5-3.5 hours under the conditions of adding 0.1-1.5mg / L BSA (bovine serum albumin), pH value 3-9 and 4 DEG C, thereby preparing the CLEAs; and the enzyme activity recycling rate of the cross-linked immobilized enzyme is up to more than 90%. The invention has the advantages of simple experimental operations and low cost; the obtained CLEAs have obviously higher stability than free oxalate decarboxylase; and thus, the invention can be used for developing enzyme preparations for oxalate calculus disease.

The invention discloses a preparation method of dry powder and fungus powder containing Tricholoma lobayense Heim oxalate decarboxylase and relates to the field of degradation of oxalate in stomachs and intestines of human bodies or animal bodies. The preparation method comprises the following steps of: using Tricholoma lobayense Heim or sibling fungi thereof as raw materials, culturing thalli by adopting a liquid fermentation method, regulating pH (potential of hydrogen) of fermentation solution to 2.0-5.0, inducing to produce oxalate decarboxylase, collection the fermentation solution and the thalli, crushing the thalli, conducting solid-liquid separation to respectively obtain liquid and solid containing the Tricholoma lobayense Heim oxalate decarboxylase, and respectively drying the obtained liquid and solid after separation to obtain the dry powder and the fungus powder containing the Tricholoma lobayense Heim oxalate decarboxylase. The dry powder and the fungus powder containing the Tricholoma lobayense Heim oxalate decarboxylase can be used for oxalic acid removal of food, beverage, feed and the like, can also be used as raw materials to prepare healthcare products or medicines, is suitable for oral taking, can effectively degrade oxalic acid and oxalate and can prevent or cure urine oxalic acid excess symptoms and calcium oxalate calculi.

The invention discloses a preparation method of oxalate decarboxylase. The method comprises the following steps: constructing a recombinant expression plasmid vector comprising Bacillus-containing acid induced promoter, a secretion signal peptide, and a fungus-derived oxalate decarboxylase gene prepared under the control of the secretion signal peptide, transforming bacilli normally growing in a pH value being not more than 4.5 by the vector to prepare oxalate decarboxylase recombinant bacteria; and carrying out culture and acid feeding on the recombinant bacteria in a fermentation medium, inducing the recombinant bacteria, and carrying out simple purification to recover generated extracellular oxalate decarboxylase. The extracellular oxalate decarboxylase prepared through the method is used in diagnosis reagent raw materials, medicinal compositions, healthcare foods or formulated foods with special medical uses. The mutated acid induced promoter is adopted to substitute an original acid induced promoter in the recombinant expression vector, so the transformed bacilli have a high expression level.

The invention discloses a fresh polygonatum odoratum freeze-dried decoction piece and a preparation method thereof, and relates to the field of traditional Chinese medicine, the fresh polygonatum odoratum is cleaned to remove impurities, the cleaned fresh polygonatum odoratum is cut into thick pieces, the thick pieces are placed in an acidic soak solution containing vegetable oil for ultrasonic impregnation, and the thick pieces are taken out and dried to obtain a first crude product; the first crude product is put into fermentation liquor containing oxalate decarboxylase to be fermented, and a second crude product is obtained after fermentation is finished; the second crude product is subjected to freeze drying under the vacuum condition, and a third crude product is prepared. According to the fresh polygonatum odoratum freeze-dried decoction piece, the content of calcium oxalate in the fresh polygonatum odoratum freeze-dried decoction piece is reduced, and meanwhile the dried decoction piece is not prone to embrittlement and easy to store.

The invention relates to a method for culturing coriolus versicolor and inducing oxalate decarboxylase by a rice straw carbon source. The method comprises the following steps of: (1) dipping pulverized rice straws by using an acid solution and hydrolyzing under the condition of 100-150 DEG C; (2) preparing a fermentation culture medium and regulating the pH value to be 4.0-5.0; (3) inoculating mycelia into the liquid seed culture medium, transferring both the mycelia and the culture medium into a container containing sterile glass beads when a white mycelium group is formed but does not emerge from the water level, shaking, smashing and inoculating the mycelia into the fermentation culture medium according to an inoculating quantity of 3-15 percent; (4) culturing for 4-6 days, adding oxalic acid and then culturing 0.5-6 days to obtain the mycelia; and freezing the mycelia by liquid nitrogen, then grinding, smashing, extracting mycelium fragments by an acetic acid buffer solution and centrifuging. The method is simple and easy to operate, and has low cost, strong controllability of carbon source preparation of and stable sugar content.

The invention relates to a method for modifying oxalate decarboxylase, particularly a method for modifying oxalate decarboxylase with monomethoxypolyethylene glycol, belonging to the technical field of biochemical engineering. The method comprises the following steps: preparation of recombinant oxalate decarboxylase, activation of monomethoxypolyethylene glycol (mPEG), modification of oxalate decarboxylase and the like. The modified oxalate decarboxylase has higher heat stability, pH adaptability, heat resistance and trypsinase tolerance. The method solves the realistic problem that the oxalate decarboxylase in use can be influenced by digestion of gastric acid and proteinase and can be easily digested to lose activity at present. The method can be used for preparing an enzyme preparation for preventing and treating calculus caused by oxalate crystallization.

The invention discloses a recombinant oxalate decarboxylase expressed by a filamentous fungal host cell, a construction method of the recombinant bacterial strain, a production method and application of the recombinant enzyme, and belongs to the technical field of genetic engineering. The recombinant filamentous fungal host cell comprises one or more copies of an oxalate decarboxylase expression cassette integrated in its genome, the oxalate decarboxylase expression cassette including a promoter, a signal peptide coding sequence, an oxalate decarboxylase coding gene and a terminator. The above-mentioned host cells can be constructed by random integration or site-specific integration. In addition, the present invention also optimizes the medium formulations for different recombinant filamentous fungal host cells. The production of oxalate decarboxylase, through the construction of expression cassettes, vector construction, host cell construction and the adjustment of the final culture positive formula, the final yield and enzyme activity are greatly improved, effectively solving the problem of oxalate decarboxylase in the prior art. Artificial production cannot be industrialized on a large scale.

The invention discloses a method for modifying oxalate decarboxylase by means of ethylenediaminetetraacetic dianhydride. The method comprises the steps of dissolving ethylenediaminetetraacetic dianhydride and oxalate decarboxylase in phosphate buffer, and conducting stirring reaction for 1-24 h; dialyzing the reaction product after reaction ends, and then conducting concentration to obtain ethylenediaminetetraacetic dianhydride modified oxalate decarboxylase. Chemical modification of oxalate decarboxylase is directly conducted with ethylenediaminetetraacetic dianhydride, conditions are mild, operation is easy, and cost is low; the heat stability, heat resistance, pH adaptability and trypsin tolerance of modified oxalate decarboxylase are improved; the adsorption capacity of oxalate decarboxylase on calcium oxalate crystals is improved especially; the problem that oxalate decarboxylase can be affected by gastric acid and pepsin in use to lose activity easily is solved; the practical problem that zymoprotein in enzymic preparations is diluted easily to disappear in practical use is also solved.