28 results about "Dehalogenase" patented technology

The invention discloses a haloalkane dehalogenase mutant and an application thereof in degradation of 1, 2, 3-tribromopropane, haloalkane dehalogenase DhaA31 coded by dhaA gene of a Rhodococcus rhodococcus NCIMB 13064 strain is modified to obtain the haloalkane dehalogenase mutant F168W / I246C, the haloalkane dehalogenase mutant has high degradation efficiency on haloalkane pollutant 1, 2, 3-tribromopropane, and the haloalkane dehalogenase mutant has high degradation efficiency on haloalkane pollutant 1, 2, 3-tribromopropane under the condition of water phase normal temperature. The degradation rate of the alkyl halide dehalogenase mutant to 10 mM of 1, 2, 3-tribromopropane within 1 h reaches 100%, and the degradation rate of the alkyl halide dehalogenase mutant to 10 mM of 1, 2, 3-tribromopropane reaches 100%. The haloalkane dehalogenase mutant has high stereoselectivity on (R)-2, 3-dibromo-1-propanol, and the e.r. Value of the haloalkane dehalogenase mutant reaches 6: 94. The haloalkane dehalogenase mutant provided by the invention has high catalytic efficiency and high stereoselectivity, is beneficial to recognition and further degradation of downstream enzymes, and has a good application prospect in bioremediation of emerging haloalkane pollutants.

The invention provides a halohydrin dehalogenase mutant and an application of the halohydrin dehalogenase mutant in synthesis of a chiral gamma-amino alcohol compound. Halogenated alcohol dehalogenase mutants enabling 2-substituted oxetane to be subjected to ring opening are obtained through the technologies of gene mining, directed evolution and the like, and the enzymes can directly perform ring opening on a substrate 2-aryl oxetane under the action of a nucleophilic reagent to generate a series of chiral gamma-amino alcohol compounds which are important precursors for synthesizing drugs. Therefore, the halohydrin dehalogenase mutants have important application value in the industry.

The present invention discloses a haloalkane dehalogenase and its application in the degradation of 1,2,3-trichloropropane. The haloalkane dehalogenase provided by the present invention is a protein with the amino acid sequence of SEQ ID NO.1. The haloalkane dehalogenase of the present invention has high degradation activity towards the refractory and highly toxic 1,2,3-trichloropropane, and can degrade 42.63% of 1,2,3-trichloropropane after 2 h of reaction, with a fast biodegradation rate; IPTG is used to induce the expression of the haloalkane dehalogenase in recombinant bacteria, and high enzyme activity can be obtained after 5 h of expression. The recombinant bacteria express the haloalkane dehalogenase faster and with higher yields. The haloalkane dehalogenase of the present invention has high activity in a relatively wide pH range (3.0 - 9.0) and temperature range (25°C - 60°C), and has a relatively broad application scope.

The invention discloses a gas-phase white carbon black tail gas dechlorination process, which belongs to the technical field of tail gas treatment, and comprises tail gas pretreatment and particulate matter separation, germanium molecular sieve carrier preparation and modification, Fe-Cu bimetallic nano-cluster loading, dynamic pulsed electric field activation, dry-process dechlorination, wet-process reduction synergistic dechlorination and photo-thermal coupling regeneration. According to the method, the alkyl halide dehalogenase is introduced as a stabilizer, and the Fe3O4-CuO nano-cluster activated by the dynamic pulsed electric field is adopted, so that the reaction process is remarkably optimized, the alkyl halide dehalogenase and active components in the sodium thiosulfate solution form synergistic action sites, the selectivity and efficiency of sodium thiosulfate participating in the reaction are improved, and the reaction time is shortened. The consumption in a non-target reaction is avoided; according to the Fe3O4-CuO nano-cluster activated by the dynamic pulsed electric field, the chlorine removal efficiency is improved, and the usage amount of sodium thiosulfate is reduced, so that the consumption amount of sodium thiosulfate and the generation amount of waste liquid are synchronously reduced, and the high-cost investment of enterprises on waste liquid treatment and the risk of environmental protection compliance are reduced.

The invention discloses a haloalkane dehalogenase mutant with high stereoselectivity and application thereof.The haloalkane dehalogenase mutant F131M / F168M / I246L is obtained by modifying haloalkane dehalogenase DhaA31 coded by a rhodococcus dhaA gene, the haloalkane dehalogenase mutant has high degradation efficiency on haloalkane pollutants 1, 2, 3-tribromopropane, and the haloalkane dehalogenase mutant has high stereoselectivity on haloalkane pollutants 1, 2, 3-tribromopropane under the water-phase normal-temperature condition. The degradation rate of the alkyl halide dehalogenase mutant to 10 mM of 1, 2, 3-tribromopropane within 60 min reaches 100%, and the degradation rate of the alkyl halide dehalogenase mutant to 10 mM of 1, 2, 3-tribromopropane reaches 100%. The haloalkane dehalogenase mutant has high stereoselectivity on a high-value product (S)-2, 3-dibromo-1-propanol, and the e.r. Value of the haloalkane dehalogenase mutant reaches 98: 2. The haloalkane dehalogenase mutant provided by the invention has high catalytic efficiency and high stereoselectivity, and has a good application prospect in bioremediation and recycling of emerging haloalkane pollutants.

The invention relates to the technical field of biology, in particular to a method for increasing the growth rate of a Cupriavidus hookworm strain with carbon dioxide as a carbon source and an engineering strain. Glucokinase genes participating in glucose metabolism in the Cupriavidus hookworm strain are knocked out, alkaline phosphatase genes and haloid acid dehalogenase family (HAD) phosphatase genes are knocked out on the basis, and the growth rate of the Cupriavidus hookworm strain under the condition that carbon dioxide serves as a carbon source is increased through the operation. The bacterial strain with the increased proliferation rate can be used for producing single-cell protein by taking carbon dioxide as a raw material and is beneficial to fixation of greenhouse gas carbon dioxide. Meanwhile, the production efficiency can be remarkably improved by utilizing the chassis cell to construct a metabolic engineering-based product synthesis strain.

The present invention relates to the field of TREM2 receptor drug screening, and specifically proposes a rapid screening method for TREM2 receptor drugs. The method mainly comprises: (1) fusing a halogenated alkane dehalogenase to the inactive end of the TREM2 receptor to construct a TREM2 recombinant protein with a directional immobilization function; (2) covalently modifying a separation medium with 6-bromohexanoic acid to obtain a surface-modified separation medium; (3) fully mixing the TREM2 recombinant protein with the 6-bromohexanoic acid-modified separation medium, and incubating the mixture to allow the TREM2 receptor to be directionally modified on the surface of the separation medium. The resulting TREM2 recombinant protein immobilized complex is further used to prepare a TREM2 receptor chromatographic column, which can specifically identify its ligand based on retention time, thereby realizing rapid screening of TREM2 receptor-targeting compounds in complex systems such as traditional Chinese medicine.

The invention provides a method for improving the stability of dehalogenase through an arabinogalactan covalent cross-linking immobilization technology. The method comprises the steps of constructing and expressing a recombinant dehalogenase strain, purifying the recombinant dehalogenase and carrying out dehalogenase covalent cross-linking immobilization. The arabinogalactan is used for covalently crosslinking the dehalogenase, so that the thermal stability of the dehalogenase and the stability of the dehalogenase under different pH conditions and in an organic solvent can be remarkably improved.

The invention discloses halogenated alkane dehalogenase HLD-1405 as well as a coding gene and application thereof, and belongs to the technical field of biology. The amino acid sequence of the halogenated alkane dehalogenase HLD-1405 is as shown in SEQ ID NO. 2, and the coding gene of the halogenated alkane dehalogenase HLD-1405 is as shown in SEQ ID NO. 1. The halogenated alkane dehalogenase HLD-1405 has high catalytic activity on 1, 3-dibromopropane, the optimum action condition is 40 DEG C / pH 9.0, and the activity of the halogenated alkane dehalogenase HLD-1405 in 20% methanol is kept 100%. The haloalkane dehalogenase HLD-1405 can efficiently degrade short-chain haloalkane, has the capability of degrading pollutants such as DDT and hexachlorocyclohexane, has great application potential in treatment of stubborn pollutants, and is suitable for bioremediation and industrial catalysis of environmental pollution.

This invention discloses a haloalkane dehalogenase HLD-1405, its encoding gene, and its applications, belonging to the field of biotechnology. The amino acid sequence of this haloalkane dehalogenase HLD-1405 is shown in SEQ ID NO.2, and its encoding gene is shown in SEQ ID NO.1. This haloalkane dehalogenase HLD-1405 exhibits high catalytic activity towards 1,3-dibromopropane, with optimal operating conditions of 40℃ / pH 9.0, and retains 100% activity in 20% methanol. This haloalkane dehalogenase HLD-1405 can efficiently degrade short-chain haloalkanes and also has the ability to degrade pollutants such as DDT and hexachlorocyclohexane (HCH), showing great application potential in the treatment of stubborn pollutants and suitable for environmental bioremediation and industrial catalysis.

The invention belongs to the technical field of enzyme engineering, and particularly relates to application of halogenated alkane dehalogenase in selective preparation of S-configuration aryl ethanol, the halogenated alkane dehalogenase is DbjA, and the amino acid sequence of the halogenated alkane dehalogenase is shown as SEQ ID NO.1. According to the invention, the halogenated alkane dehalogenase DbjA has stereoselectivity for the reaction of catalyzing phenyl chloride to prepare S-configuration aryl ethanol, the enantiomer selectivity for the catalytic reaction of different substrates can reach 500 or more, the ee value of the generated substrate can reach 99%, and meanwhile, the yield of 40% or more can be maintained; and a new solution is provided for the problems of complex operation and large workload in the preparation process of S-configuration aryl ethanol.

The invention relates to a microbial agent for degrading trichloroethane as well as an enrichment method and application of the microbial agent, and belongs to the technical field of microbial engineering. The invention provides a microbial agent for degrading trichloroethane, which comprises a functional bacterium A and a functional bacterium B. The functional bacterium A and the functional bacterium B have amino acid sequences for coding dehalogenase a and / or dehalogenase b, the amino acid sequence for coding dehalogenase a is shown as SEQ ID NO.3, and the amino acid sequence for coding dehalogenase b is shown as SEQ ID NO.4. The invention further provides a preparation method of the microbial agent for degrading trichloroethane. Experiments find that 1, 1, 2-TCA can be degraded into 1, 2-DCA by the dehalogenase a under the condition that the dehalogenase a and the dehalogenase b exist at the same time, and then the 1, 2-DCA is instantly converted into ethylene by the dehalogenase, so that only a small amount or no toxic intermediate product 1, 2-DCA or VC is generated in the degradation process.

The invention relates to the technical field of chemical genetics marking, in particular to a marking probe for quantifying protein dynamics. The probe comprises a compound with the following structural formula, wherein the compound contains a halogenated alkane chain and can be specifically covalently bound with halogenated alkane dehalogenase-HaloTag tag protein. The HaloTag-containing fusion target protein is combined with the HaloTag-containing fusion target protein in cells, and has the advantages of high specificity and stability, low toxicity, good biocompatibility and strong photobleaching resistance in living cell imaging. The compound has the property of space-limited emission effect, and can sensitively mark and respond to the spatial dynamic change of any single protein molecule in the physiological process of cells through a HaloTag bridge, so that the dynamic quantification and visualization of the protein are realized in the method, and the compound is used for recording physiological activities under different time scales.

The invention discloses preparation and application of a protein composite imprinting material for targeted recognition of strigolactones. The method comprises the following steps: 1, fusing halogenated alkane dehalogenase into ShHTL7 for induced expression to obtain recombinant protein; 2, preparing nano magnetic ferroferric oxide hollow microspheres; 3, carrying out surface amination modification on the nano magnetic ferroferric oxide hollow microspheres; 4, functionalizing halogenated alkanoic acid on the surfaces of the nano magnetic ferroferric oxide hollow microspheres subjected to amination modification; and 5, preparing the strigolactones surface protein molecularly imprinted polymer. The protein composite imprinting material which is high in adsorption efficiency, high in adsorption selectivity and capable of being rapidly enriched is prepared, shows superparamagnetism, can be rapidly separated under the action of a magnet, shows ultrahigh targeted recognition capacity on strigolactones, can be used for targeted extraction of strigolactones in parasitic plant root exudates, and has a good application prospect. The materials are convenient and fast to recycle and can be repeatedly used.

The invention discloses a method for biocatalytically synthesizing optically pure 2R-fluorocarboxylic acids and 2R-hydroxycarboxylic acids, and belongs to the field of bioengineering technology. Using α-fluoro acid and / or α-fluoro acid derivatives as substrates, using fluoroacetic acid dehalogenase mutants K181M, K181S, W185G, K181M-W185Y, K181S-W185A, K181S-W185C, K181S-W185G, K181S-W185V as biocatalysts, hydrolysis and defluorination obtain chirally pure 2R-fluorocarboxylic acids and 2R-hydroxycarboxylic acids and derivatives thereof. The present invention has strong stereoselectivity and simple steps, and solves the problems of low optical purity, low yield and environmental pollution that are widely present in the synthesis of chiral compounds. In addition, the inventive method amplifies the reaction system to gram-level experiments using whole cells as catalysts, obtains products with higher yields and ee values, has low cost, low energy consumption, and is suitable for industrial production.

The invention relates to the field of gene engineering and enzyme engineering, in particular to a halohydrin dehalogenase mutant HheC-3 (F12Y / P84T / F86W) and application of the halohydrin dehalogenase mutant HheC-3 in synthesis of chiral vicinal diol, and an amino acid sequence corresponding to the HheC-3 is shown as SEQ ID No.4. The halohydrin dehalogenase mutant can be used for catalyzing a series of racemic epoxide I or racemic o-halohydrin compound II enantiomers to polymerize into a single chiral vicinal diol compound III, the separation yield is as high as 91%, and the optical purity is as high as 96: 4 er (enantiomer ratio).

The present application relates to the technical field of chemical genetics marking, in particular to a kind of probe for marking quantifying protein dynamics.The probe includes the compound of the following structural formula: the compound contains halogenated alkane chain, can be specifically covalently combined with halogenated alkane dehalogenase-HaloTag label protein.It is combined with the fusion protein containing HaloTag in cell with the purpose of protein, with high specificity and stability, low toxicity, with good biocompatibility and strong anti-photobleaching ability in live cell imaging.The compound has the nature of space limited emission effect, through HaloTag bridge, can sensitively mark and respond to the spatial dynamic change of any single protein molecule in cell physiological process, and then realize the quantification and visualization of protein dynamics in method, for recording physiological activity under different time scales.

Disclosed herein are compounds that have improved cellular specificity. The compounds have linkers and other moieties that can both aid in cellular specificity and that can attach functional groups to a target cell. The compounds can be used, e.g., in methods of modulating, detecting, and labeling of proteins and cells. An example method includes the compound forming a covalent bond with a dehalogenase variant.

The invention relates to the technical field of genetic engineering, in particular to an engineering bacterium capable of producing 2-halogenated acid dehalogenase at high yield as well as a construction method and application of the engineering bacterium. Trichoderma viride 2-halogenated acid dehalogenase gene TvHAD1 is over-expressed in escherichia coli or pichia pastoris, engineering bacteria of the 2-halogenated acid dehalogenase are constructed, and the engineering bacteria of the high-yield 2-halogenated acid dehalogenase are obtained through collaborative optimization of fermentation processes such as induction conditions and metal ions. And efficient heterologous expression and large-scale preparation of the enzyme are realized. Under the shake flask scale, the enzyme yield of the recombinant pichia pastoris engineering bacteria can reach 102.51 mg / L, the enzyme yield of the recombinant escherichia coli engineering bacteria is 21.07 mg / L, and high yield of the 2-halogenated acid dehalogenase is realized.

The invention discloses an immobilized alkyl halide dehalogenase, a preparation method thereof, a battery system and a dehalogenation method for catalyzing hydrolysis of halogenated organic matters. The immobilized alkyl halide dehalogenase comprises a carrier and an alkyl halide dehalogenase mutant fixed on the carrier, the carrier is a conductive two-dimensional carbon material of which the surface contains carboxyl; the carbon terminal of the alkyl halide dehalogenase mutant has a cysteine residue, and the cysteine residue is the only cysteine residue on the protein surface; the carrier is connected with the alkyl halide dehalogenase mutant through a connecting agent, and end groups at two ends of the connecting agent are maleimide group and amino group respectively. According to the immobilized alkyl halide dehalogenase and the battery system, the problem of product inhibition caused by halide ion enrichment is solved, and the stability of the enzyme is improved.

This invention relates to the fields of enzyme engineering and biocatalysis, disclosing a halohydrin dehalogenase mutant and its catalytic enantioselective dehalogenation hydroxylation reaction of a series of racemic ε-halohydrins, preparing corresponding chiral ε-diols and chiral ε-halohydrins. The invention utilizes a fungicide derived from *Agrobacterium radiodurans* (…). Agrobacterium tumefaciens The wild-type haloalcohol dehalogenase HheC (amino acid sequence SEQ ID NO:2) of AD1 was used as the parent enzyme, and was obtained by combining 2 to 6 amino acid site mutations at L142, W249, P84, T134, N176, and F186. The mutant can be used to catalyze the kinetic resolution and dehalogenation reactions of racemic ε-haloalcohols, synthesizing a series of high-optical-purity chiral ε-diols and chiral ε-haloalcohols. These compounds are important chiral building blocks for the preparation of various pharmaceutical chemicals and have broad industrial application value.

The application discloses a catalytic system and method for hydrolysis of halogenated organic matter catalyzed by enzyme at an oil-water interface. The catalytic system comprises an organic solvent, an aqueous solution and an enzyme-polymer conjugate enzyme; the enzyme-polymer conjugate enzyme is a conjugate of halogenated alkane dehalogenase and amphiphilic block copolymer covalently combined; in the enzyme-polymer conjugate enzyme, 2-3 molecules of amphiphilic block copolymer are connected to each molecule of enzyme. The application enriches the enzyme at the oil-water interface by constructing an oil-water two-phase reaction system and a conjugate synthesized by the amphiphilic block polymer and the enzyme; the accessibility of the substrate in the organic solvent is improved, meanwhile, the enzyme inactivation caused by the organic solvent is avoided, and the hydrolysis activity of the enzyme on the halogenated organic matter is improved; the enzyme catalytic reaction product hydrogen halide can be dissolved in the aqueous solution by constructing the oil-water two-phase reaction system, and the reduction of the enzyme catalytic activity caused by the retention of halogen ions in the enzyme product pocket is reduced.

The application discloses an ancestral dehalogenase Ans171 and application thereof in degrading sulfur mustard halogenated pollutants. The amino acid sequence of the ancestral dehalogenase Ans171 is shown as SEQ ID NO. 2, and the nucleotide sequence of the coding gene is shown as SEQ ID NO. 3. The ancestral dehalogenase Ans171 of the application can efficiently degrade sulfur mustard simulant agent dichlorodiethyl sulfide. After 200 μL of crude enzyme solution of the ancestral dehalogenase Ans171 is added into 2 μL of dichlorodiethyl sulfide and reacted for 5 min, the degradation rate reaches 99.7%. The thermal stability and degradation efficiency of the ancestral dehalogenase Ans171 of the application are significantly higher than those of a modern enzyme, and the ancestral dehalogenase Ans171 has a good application prospect in bioremediation of sulfur mustard halogenated pollutants.

The invention relates to the technical field of enzyme engineering, namely biological catalysis, and discloses a halohydrin dehalogenase mutant and application of the halohydrin dehalogenase mutant to catalysis of enantioselective desymmetry of a series of 3-substituted oxetane and synthesis of corresponding chiral gamma-azido alcohol. Wild halohydrin dehalogenase (the amino acid sequence is SEQ ID NO: 2) derived from Ilumacter coccineus YM16-304 is used as a parent enzyme, and 2-5 amino acid site combination mutation is carried out on sites I104, Y18, T106, V105 and L103, so that the halohydrin dehalogenase is obtained. The chiral gamma-azido-alcohol is applied to catalysis of enantioselective azidation reaction of 3-substituted oxetane and sodium azide to synthesize a series of chiral gamma-azido-alcohol with high optical purity, and can be used as a key chiral intermediate to prepare chiral blocks such as triazole compounds, gamma-amino-alcohol and macrolactam. The method has application value in preparation of various bioactive drugs and fine chemicals.

The application relates to the technical field of inks, in particular to a high-flexibility acrylic epoxy resin for inks and a preparation method thereof. The preparation method of the high-flexibility acrylic epoxy resin for inks comprises the following steps: synthesizing an acrylic copolymer, carrying out a catalytic reaction of immobilized halogen alcohol dehalogenase, adding a flexible modifier, catalyzing by immobilized lipase and carrying out a grafting reaction of an epoxy resin. The application introduces long flexible side chains by enzyme-catalyzed open-loop grafting of flexible segments, improves the activity of molecular segments on the basis of retaining the original reaction activity and skeleton strength of the acrylic epoxy resin, improves the packing density of the long flexible side chains, improves the activity of molecular segments, and when the prepared acrylic epoxy resin is used in inks, the flexibility of ink films can be improved, the curing speed and surface hardness can be considered, meanwhile, the acrylic epoxy resin does not cause adverse effects on the transparency and storage stability of the ink films.

The invention discloses a halohydrin dehalogenase directional modification method for synthesis of an atorvastatin intermediate, and belongs to the technical field of enzyme directional modification, and the method comprises the following steps: downloading an amino acid sequence of H halohydrin dehalogenase from an NCBI (National Center of Biotechnology Information) protein database, carrying out multi-sequence alignment by using CLUSTALW software, carrying out molecular dynamics simulation, then carrying out PEGylation modification, and carrying out enzyme directional modification. The directionally modified halohydrin dehalogenase is prepared. According to the immobilization method combining the directionally modified halohydrin dehalogenase and the functionalized magnetic porous carrier, the intrinsic activity and stability of the enzyme are improved through directional modification, and the functionalized carrier provides a suitable microenvironment for the enzyme and realizes firm immobilization; the invention not only solves the technical problems of large activity loss, poor stability and the like in the traditional enzyme immobilization process, but also provides an efficient, stable and reusable biocatalyst for industrial application.

The invention belongs to the technical field of biology, and particularly relates to a dehalogenase HcdA as well as a coding gene and application thereof. The amino acid sequence of the dehalogenase is SEQ ID NO: 2. The dehalogenase shows excellent degradation capability on 3-chloro-2-methyl propylene and 3-bromine-2-methyl propylene under wide temperature conditions and pH conditions, and has the potential of biological remediation of the ecological environment.