191 results about "Oxygenase" patented technology

The present invention provides a 4-vinylguaiacol oxygenase mutant for synthesizing vanillin, and an engineered bacterium thereof. The 4-vinylguaiacol oxygenase mutant is an enzyme having the sequence as shown in SEQ ID NO. 1, which exhibits a double-site mutation of D27P / T216M, D27P / M351F, D27P / Y156D, M351F / Y156D, T216M / Y156D or T216M / M351F, or a single-site mutation of T216M, T216L, S280M, G307P or M351F, and can be used for catalyzing 4-vinylguaiacol to generate vanillin.

The invention discloses a Vinck Cellulomicrobe and application thereof in degradation of corn straw and improvement of soil fertility, and belongs to the field of agricultural microbial technology and solid waste resource utilization. The bacterial strain can secrete a plurality of hydrolase and oxidase including AA10 family lytic polysaccharide monooxygenase, AA2 peroxidase, GH3 and GH1 beta-glucosidase, GH16 / GH17 beta-glucosan endonuclease and AA1 multi-copper oxidase. The bacterial strain has the advantages that the bacterial strain can secrete a plurality of hydrolase and oxidase; fermentation liquor is applied to soil according to the straw mass ratio of 1: 5, and the corn straw degradation rate can be increased under the condition that the mass ratio of soil to straw is 100: 1. Meanwhile, the microbial agent can synergistically improve soil nutrients and increase the contents of rapidly available phosphorus, rapidly available potassium, alkali-hydrolyzable nitrogen, soluble organic carbon and total phosphorus in soil. According to the microbial agent, a straw lignocellulose structure is effectively cracked through a multi-enzyme system synergistic effect, and the dual effects of accelerating straw degradation and improving soil fertility are achieved.

The invention relates to the technical field of carotenoid cleavage dioxygenase, and discloses a carotenoid cleavage dioxygenase mutant and application thereof. In order to improve the efficiency of converting carotenoid in tobacco into aroma molecule beta-ionone, specific mutation of characteristic sites is carried out on carotenoid cleavage dioxygenase (PhCCD1) from petunia, that is, serine (S) at the 428 site of the carotenoid cleavage dioxygenase is mutated into cysteine (C) or tyrosine (Y) to obtain a mutant S428C or S428Y, and the mutant S428C or S428Y is converted into beta-ionone. The yield of the beta-ionone converted from the carotenoid is greatly improved. Experiments prove that the yields of beta-ionone of S428C and S428Y respectively reach 54.2 mg / L and 50.6 mg / L, which are respectively increased by 69.7% and 58.5% compared with those of a wild strain WT.

The invention discloses a heme oxygenase mutant and application of the heme oxygenase mutant in preparation of bilirubin through multienzyme coupling, and belongs to the field of biological catalysis engineering. The forward mutant of the heme oxygenase is obtained through a directed evolution technology, and heme can be efficiently catalyzed to synthesize biliverdin; the bilirubin is synthesized by further introducing biliverdin reductase, coupling cytochrome P450 reductase and formate dehydrogenase, and coupling the two core enzymes with foreign assistant enzymes to construct recombinant escherichia coli to catalyze heme to synthesize bilirubin. In the system, NADPH does not need to be exogenously added, the reaction time is about 4 h, the yield of bilirubin reaches 2.10 g / L, accumulation of intermediate products is not observed in the catalysis process, and the highest yield and the shortest reaction time in bilirubin synthesis reported at home and abroad at present are achieved. Compared with the traditional pig bile extraction, the method disclosed by the invention breaks through the resource limitation, is simple to operate and high in yield, and realizes efficient and green synthesis of bilirubin.

Described herein are biocatalytic methods of producing terpene degradation products useful as starting material for the production of perfumery ingredients, such as, for example, ambrox. In particular novel terpene degrading polypeptides (enal-cleaving polypeptides) and novel peptides converting terpenes compounds to oxygenated derivatives (oxygenases) and mutants and variants derived therefrom are described which may be applied in novel types of fully enzymatic multistep degradation pathways allowing the controlled, stepwise conversion and degradation of linear or cyclic terpene substrates. Said novel biosynthetic strategies allow the fully biochemical synthesis of valuable terpene-derived compounds, like for example manooloxy or gamma ambrol. Also described herein are recombinant host organisms carrying the required set of genetic information for the functional expression of the set of enzymes necessary for catalyzing the combination of enzymatic conversion and degradation steps.

The invention discloses an alcohol dehydrogenase mutant for high yield of tea flavor ketone and application of the alcohol dehydrogenase mutant in a one-pot method. The mutant is A182L or A182L / V214T. The alcohol dehydrogenase mutant has high catalytic activity on 4-hydroxyisophorone, and the yield of tea flavor ketone prepared by catalysis of the mutant A182L / V214T can reach 97% and is 3.73 times that of an original enzyme; the alcohol dehydrogenase mutant has high catalytic activity of synthesizing the tea aroma ketone through one-pot catalysis of isophorone, and the yield of the tea aroma ketone obtained through direct oxidation of isophorone by jointly using the mutant A182L / V214T and a cytochrome P450BM3 monooxygenase mutant can reach 92%; the alcohol dehydrogenase mutant has the advantages of simple production process, mild reaction conditions and environment-friendly production process, is beneficial to the industrial production of the tea flavor ketone, and has wide application prospects.

The invention discloses a preparation method of recombinant catechol-2, 3-dioxygenase and an application of the recombinant catechol-2, 3-dioxygenase. According to the invention, a novel catechol-2, 3-dioxygenase group C23O927 is identified from a constructed metagenome library, and the catechol-2, 3-dioxygenase group C23O927 is a key enzyme for catalyzing degradation of polycyclic aromatic hydrocarbon pollutants. A C23O927-pET32a (+) recombinant expression vector is constructed and converted into host bacteria, heterologous expression of target protein is achieved, and the prepared recombinase belongs to acid enzymes and has good resistance to high temperature, acid, alkali, high-concentration metal ions, organic solvents and surfactants. The recombinase provided by the invention can be used in an acidic environment, has very good enzymatic activity, can catalytically degrade various catechol compounds, can be better used for treating polycyclic aromatic hydrocarbon polluted environments, and has important significance for developing more products which can tolerate various extreme environments and efficiently catalyze degradation of polycyclic aromatic hydrocarbons.

The invention relates to a method for increasing the yield of glucose biologically converted from corn straws, which comprises the following steps of: carrying out composite pretreatment on the corn straws by using NaOH and urea, and then carrying out step-by-step enzymolysis, namely carrying out enzymolysis on the corn straws by using pleurotus eryngii lysate polysaccharide monooxygenase in the first step, and adding cellulase for further enzymolysis to produce the glucose in the second step, the pleuromutilus rubra lysate polysaccharide monooxygenase is a compound enzyme composed of PdLPMO9A and PdLPMO9B according to a ratio of (1: 2)-(1: 4). According to the method disclosed by the invention, the corn straws are subjected to specific pretreatment, the compound enzyme is formed by adopting the pleurotus eryngii lysate polysaccharide monooxygenase PdLPMO9A and PdLPMO9B according to a specific proportion, and the compound enzyme and the cellulase are used for performing enzymolysis on the corn straws step by step, so that the biological conversion efficiency of the corn straws is effectively improved under the same enzymolysis time, and the yield of glucose is improved and reaches 0.642 g / g of straws.

The present application relates to a kind of sulfide monooxygenase mutant and its application in esomeprazole synthesis.The sulfide monooxygenase mutant, its encoding gene, the preparation method of the recombinant expression vector containing the gene sequence, co-expression recombinant vector and recombinant expression transformant, and the application of the recombinant sulfide monooxygenase mutant catalyst in esomeprazole synthesis are specifically disclosed.Compared with other sulfide monooxygenases, the sulfide monooxygenase mutant obtained by the present application can efficiently utilize the coenzyme NADH with lower cost and higher stability to catalyze the asymmetric oxidation reaction of omeprazole sulfide, prepare esomeprazole, has the advantages of high substrate concentration, mild reaction condition, environment-friendly, simple operation, high yield, low production cost and good industrial application prospect.

The invention relates to a yarrowia lipolytica strain for producing tyrosol as well as a construction method and application of the yarrowia lipolytica strain, and belongs to the technical field of genetic engineering. According to the present invention, ScADH6, ScARO10F138L and D218G genes are expressed in the yarrowia lipolytica W29 [delta] KU70; the shikimic acid feedback inhibition is relieved by expressing the Y1ARO3K225L gene, the Y1ARO4K2214 gene and the Y1AR07G139S gene; pmLAAD, EcTyrAM53I and A354V genes are introduced, so that the conversion from FAD to FADH2 is promoted, and the supply of NADH (Nicotinamide Adenine Dinucleotide Hormone) in cytoplasm is enhanced; a 4-hydroxyphenylpyruvate dioxygenase (HPD) gene is knocked out; and overexpressing Y1DHS1, Y1DHS2 and Y1DHS3 genes of a shikimic acid pathway to obtain the yarrowia lipolytica strain HY5 for producing tyrosol. The invention lays a foundation for industrial production of tyrosol, and has important social significance and economic value.

The invention discloses a cephalotaxus hainanensis Ch2OGD1 gene and application thereof, and belongs to the technical field of gene engineering. The Ch2OGD1 gene is used for coding a 2-oxoglutaric acid dependent dioxygenase (2OGD) type Pictet-Spengler enzyme, and the Pictet-Spengler enzyme is used for coding a Pictet-Spengler enzyme. Through a tobacco transient expression system and in-vitro enzyme activity determination, the Ch2OGD1 is proved to be capable of specifically catalyzing the condensation of dopamine and 4-hydroxyphenylpropanal to generate 1-phenethyl isoquinoline. The invention provides a directional preparation method of 1-phenethyl isoquinoline based on the Ch2OGD1 gene, provides a key enzyme element for biosynthesis of a cephalotaxine alkaloid precursor compound, and provides a technical basis for expanding the application of the gene in plant genetic engineering. And a potential technical path is provided for relieving the problem of resource shortage of cephalotaxine alkaloids.

A genetically modified yeast cell, wherein the yeast cell comprises a genetic modification comprising overexpression of yeast gene encoding porphobilinogen deaminase (HEM3), the HEM3 gene having at least 80% identity with SEQ ID No. 7. The genome of the modified yeast cell further comprises one or more genetic modifications in one or more genes selected from: genes coding for heme-dependent repressor of hypoxic genes (ROX1), genes coding for heme oxygenase (HMX1), genes coding for a receptor for vacuolar proteases (VPS10), and genes coding for vacuolar proteinase (PEP4), the one or more genetic modifications being such that expression of a polypeptide from such a gene is reduced or disrupted or the polypeptide expressed is non-functional.

A Levodopa-selective sensor is provided. A continuous levodopa monitor is also provided. The levodopa sensor includes a sensor area having a working electrode with a levodopa-selective chemistry configured for at least partial implantation in a host. The sensor also includes at least one membrane adjacent the levodopa-selective chemistry. The at least one membrane includes an enzyme domain that at least one enzyme selected from a tyrosinase; a mutated tyrosinase with specificity towards I-3,4-dihydroxyphenylalanine; a dihydroxyphenylalanine 4,5-dioxygenase; a mutated dihydroxyphenylalanine 4,5-dioxygenase with specificity towards I-3,4-dihydroxyphenylalanine; and combinations thereof; a synthase enzyme with specificity towards I-3,4-dihydroxyphenylalanine; a mutated synthase enzyme with specificity towards I-3,4-dihydroxyphenylalanine; a 3,4-dihydroxyphenyl-acetaldehyde synthase; a mutated 3,4- dihydroxyphenyl-acetaldehyde synthase with specificity towards I-3,4-dihydroxyphenylalanine; a dioxygenase enzyme with specificity towards I-3,4-dihydroxyphenylalanine; and a mutated dioxygenase enzyme with specificity towards I-3,4-dihydroxyphenylalanine.

1, 4-disubstituted-1, 2, 3 triazole compounds of formula (I): (I) capable of inhibiting heme oxygenase activity, their use as a medicament for the treatment and / or prophylaxis of pathological conditions associated with alteration of heme oxygenase activity and pharmaceutical compositions comprising the same.

The application discloses a kind of generation (4S) 4-hydroxyisophorone monooxygenase mutant and preparation method thereof, the monooxygenase is cytochrome P450 BM3 Monooxygenase from bacillus megaterium. The cytochrome P450 BM3 Monooxygenase mutant has high stereoselectivity and high reaction activity to catalyze isophorone hydroxylation, mutant R47L / Y51F / F81W / A191T / N239H / I259V / A276T / A328I / A330S / L353I / I401L / L437LL catalyzes isophorone to obtain (4S) 4-hydroxyisophorone diastereomeric excess rate can reach 95%, yield can reach 96%; the cytochrome P450 BM3 Monooxygenase mutant catalyzes isophorone synthesis (4S) 4-hydroxyisophorone process is simple, environmental protection, and industrialization prospect is good.

The invention discloses 10-hydroxydecanoic acid as well as a preparation method and application thereof, and belongs to the technical field of cosmetics, Candida lipolytica is taken as a fermentation thallus, the Candida lipolytica contains alkane monooxygenase (CYP450), alcohol dehydrogenase (ADH), fatty alcohol oxidase (FAO) and fatty aldehyde dehydrogenase (FALDH), amplification and activation are performed on the Candida lipolytica in a seed culture medium, and the Candida lipolytica is obtained. The method comprises the following steps: inoculating a seed solution into a liquid culture medium, enabling a strain to adapt to a culture environment in advance, reducing strain death caused by environmental mutation after inoculation, inoculating the seed solution into the liquid culture medium, culturing until OD600 is 45-55, realizing high-density enrichment of the strain, being beneficial to increasing the content of CYP450, ADH, FAO and FALDH, adding decane and Tween 80, and enabling the Tween 80 to be capable of increasing the permeability of a cell membrane and increasing the reaction rate, the CYP450 is used for catalyzing decane to carry out hydroxylation, and ADH, FAO and FALDH are used for carrying out oxidation, so that the high-purity 10-hydroxydecanoic acid is obtained.

The invention discloses 4-hydroxy-3-methyl benzoic acid monooxygenase, a strain and a preparation method and application thereof, and relates to the technical field of bioengineering, the nucleotide sequence of the 4-hydroxy-3-methyl benzoic acid monooxygenase is shown as SEQ ID NO: 1, and the amino acid sequence of the 4-hydroxy-3-methyl benzoic acid monooxygenase is shown as SEQ ID NO: 2 and / or SEQ ID NO: 3. The gene of the 4-hydroxy-3-methyl benzoic acid monooxygenase disclosed by the invention can be used for encoding to obtain the 4-hydroxy-3-methyl benzoic acid monooxygenase, and the obtained 4-hydroxy-3-methyl benzoic acid monooxygenase can be used for effectively catalyzing continuous oxidation of ortho-methyl of hydroxyl in 4-hydroxy-3-methyl benzoic acid; the selective continuous oxidation of the ortho-position methyl of the 4-hydroxy-3-methylbenzoic acid can be realized, the 4-hydroxyisophthalic acid is generated, and the compound 4-hydroxyisophthalic acid with industrial added value is synthesized while the selective oxidation of the methyl is solved.

The invention relates to the technical field of genetic engineering, and discloses a genetically engineered bacterium for producing beta-ionone at high yield and application of the genetically engineered bacterium. A synthetic biological plasmid modularization technology is adopted, coding genes of carotenoid cleavage dioxygenase NtCCD1 in tobacco are excavated, and specific soluble protein tags including a thioredoxin tag, a maltose binding protein tag and a glutathione transferase tag are screened, so that the content of the carotenoid cleavage dioxygenase NtCCD1 in the tobacco is determined. The NtCCD1 is subjected to single-point mutation modification of mutation of 431st-site histidine into tyrosine, lysine or glutamine, and the fermentation enzyme and the gene engineering bacterium which can efficiently catalyze splitting decomposition of beta-carotene in tobacco to produce beta-ionone are constructed. The biosynthesis of the tobacco aroma component beta-ionone is realized, and the method has high efficiency.

The application discloses a method for promoting transport and improving retinal synthesis, which is obtained by introducing an exogenous ABC transport protein Pdr5 into a host fungus and adding a copy of beta-carotene-15, 15'-monooxygenase BLH; wherein the host fungus is a genetically modified Pichia pastoris strain PP-BCCIV. The application further verifies the retinal production performance of the recombinant strain by screening different surfactants, and further improves the ability of Pichia pastoris to produce retinal. At the same time, all the retinal is extracted into the extracellular, and all the carotenes are left in the intracellular, so as to achieve the co-production of carotenes and retinal. The recombinant Pichia pastoris of the application has a simple construction method, can better promote the synthesis of retinal and other vitamin A, and can make the engineering strain utilize glucose and methanol to efficiently synthesize retinal through a 5 L fermentation tank, which is beneficial to industrial production.

The invention relates to the field of genetic engineering and fermentation engineering, and discloses a genetically engineered bacterium for producing melatonin and application of the genetically engineered bacterium in fermentation production of melatonin. According to the invention, genes of mtrA, PTPS, SPR, PCD and DHPR of related enzymes synthesized by coding 2-amino-6-(1, 2-dihydroxypropyl)-5, 6, 7, 8-tetrahydro-4 (1H)-pterin diketone and genes of L-2-amino-3 (beta-indole) propionic acid monooxygenase, oxymethyltransferase, N-acetyltransferase and decarboxylase are co-expressed on the genetically engineered bacterium, so that the gene engineering bacterium is obtained. The strain can be used for fermentation production of melatonin, and experiments show that the yield of the strain can reach 6480 mg / L.

The invention belongs to the technical field of biology, and particularly relates to a pentachloronitrobenzene rapid detection method based on enzyme catalysis reaction, which comprises the following steps: converting pentachloronitrobenzene into pentachlorophenol and nitrate radicals through an enzyme system consisting of iron thioflavin protein reductase, ferredoxin and nitrobenzene dioxygenase; and then nitrate radicals are specifically detected by a Griess reagent method so as to indirectly reflect quintozene. The detection method disclosed by the invention is good in specificity, short in detection time and remarkable in visual detection effect, and can be used for on-site rapid detection of quintozene.

Methods for producing organosulfur compounds from a prokaryotic cell comprising enzymes involved in the production of cysteamine, hypotaurine, and taurine. Said prokaryote may comprise a vanin selected from vanin-1 (vnn-1), vanin-2 (vnn-2), or vanin- 3 (vnn-3), a cysteamine dioxygenase (ado), and / or a flavin-containing monooxygenase 1 (fmo1). Methods include the expression of organosulfur compounds in Corynebacterium glutamicum with incubation conditions to promote compound production.

The invention discloses a monooxygenase gene bpfC, an enzyme coded by the monooxygenase gene bpfC and application of the monooxygenase gene bpfC. The strain DN12 is separated and can degrade the BPF and grow by taking the BPF as the unique carbon source and energy. On the basis, a BPF degradation gene cluster bpf is obtained from the strain DN12 by adopting whole genome and comparative transcriptome sequencing analysis. Three different degrading enzymes (two-component oxidase BpfAB, monooxygenase BpfC and hydrolase BpfD) are found in the gene cluster and can catalyze gradual degradation of BPF, and a simple aromatic ring compound p-hydroxybenzoic acid is generated.

The invention belongs to the technical field of biological catalysis and biological engineering, and relates to a lytic polysaccharide monooxygenase expression system and application thereof. The expression system comprises an expression vector containing a lysing polysaccharide monooxygenase gene expression cassette and a host, the host is a pyrG auxotroph type aspergillus niger strain. According to the present invention, the deltaglaA gene knockout pyrG auxotrophic aspergillus niger strain is adopted as the host, and the strong promoter GlaPr is introduced to drive the expression of the cellulose activity LPMO gene, such that the interference of the endogenous glucamylase is eliminated, and the LPMO secretion efficiency is enhanced; a dynamic fermentation process is adopted, and efficient production of the LPMO protein is achieved.

The invention belongs to the technical field of biological catalysis, and discloses a non-specific peroxygenase mutant, a preparation method and application thereof, and a preparation method of calcifediol. Compared with a wild type non-specific peroxygenase (comprising an amino acid fragment with a sequence as shown in SEQ ID NO: 1), the non-specific peroxygenase mutant provided by the invention has one or more of Y47L mutation, L83F mutation and V112E mutation. The non-specific peroxygenase mutant has excellent enzyme activity, has the advantages of high conversion rate, mild reaction conditions, low cost and the like when being applied to preparation of calcifediol, and has a good industrial application prospect.

Disclosed herein is a molecule, or a pharmaceutically acceptable salt thereof, that has a formula M1DO1-L-ME3, M1DO1 is a moiety that binds to IDO1, L is a linker covalently attaching M1DO1 and ME3, and ME3 is a moiety that binds to an E3 ubiquitin ligase. Disclosed herein are also the uses of the molecule, or a pharmaceutically acceptable salt thereof, and a pharmaceutical composition comprising the same, in a method of treating cancer in a subject in need thereof.

The invention relates to gene modification, in particular to a method for improving the catalytic capacity of engineering P450 peroxygenase hydrogen peroxide and application of the method. The invention relates to a method for improving hydrogen peroxide catalytic capacity of P450 enzyme, which comprises the following steps of: identifying a channel of the P450 enzyme by using Caver Web 1.0 software by taking a heme cofactor of the P450 enzyme as a starting point; the method comprises the following steps: selecting a target channel with the channel length of 10-30, the curvature of 0-2 and the diameter of 0-3 as a target channel, then selecting amino acid of which the side chain faces the channel at an inlet or a bottleneck of the target channel as key amino acid, and then performing specific mutation on one or more key amino acids, further, at least one channel capable of improving the hydrogen peroxide catalytic capability of the P450 enzyme is obtained, so that the catalytic capability of the P450 enzyme is improved under the catalysis of hydrogen peroxide. The catalytic activity of a plurality of mutants obtained by the invention is compared with that of a wild enzyme, and the channel with high catalytic activity is an H2O2 channel. The application of the method can improve the hydrogen peroxide utilization capability of the P450 enzyme, and further improves the activity of the catalytic substrate of the P450 enzyme.

The present application provides a kind of cytochrome P450 monooxygenase gene and the protein coded by it, recombinant plasmid and application derived from ramie, the coding region sequence of the cytochrome P450 monooxygenase gene derived from ramie is as shown in SEQ ID NO:1, the amino acid sequence of the protein coded by it is as shown in SEQ ID NO:2.The present application takes ramie as research object, obtains ramie cytochrome P450 monooxygenase gene (BnMAX1) by three generations sequencing and gene annotation comparison.Longer coding sequence of the target gene is obtained by PacBio Sequel sequencing, which is simple, fast and easy to operate.The function of the gene is verified by cloning identification, bioinformatics, expression pattern and genetic transformation analysis, which shows that the gene has the effect of relieving plant cadmium toxicity.The acquisition and application of the gene provide basic theoretical support for molecular improvement of cadmium tolerance of ramie, and provide new technical means for how to improve the ecological restoration ability of ramie under heavy metal cadmium stress.

The present application belongs to the technical field of quorum quenching enzyme preparation, and particularly relates to a multi-target quorum quenching enzyme preparation and a preparation method and application thereof, such as being mainly used for inhibiting the virulence factors of Pseudomonas aeruginosa, and including acyl homoserine lactone acyltransferase AiiO protein and 3-hydroxy-4-oxoquinolone 2,4-dioxygenase AqdC protein, and the mass ratio of the two is 20:(1-25); such as being mainly used for inhibiting the virulence factors and the synthesis of biological membranes of Pseudomonas aeruginosa, and further including deferiprone, and the mass ratio of AiiO, AqdC and deferiprone is 20:(1-25):(0.695-2.78), the present application can target the Las, Rhl and PQS of Pseudomonas aeruginosa, the yield and stability of the prepared AqdC protein are both high, the two types of enzymes are used in combination after the regulation amount, and the virulence factors regulated by different quorum sensing pathways in Pseudomonas aeruginosa are effectively inhibited; the regulation amount of deferiprone is used in combination with the two types of quorum quenching enzymes, and the virulence and biological membranes of Pseudomonas aeruginosa can be effectively inhibited in a comprehensive manner.