580 results about "Stereoselectivity" patented technology

The invention provides a carbonyl reductase mutant. The carbonyl reductase mutant is obtained by performing single-point mutation or multi-point combined mutation on the 17th site, the 40th site and the 64th site of an amino acid sequence as shown in SEQ ID NO.1. The invention also provides a coding gene, a recombinant vector containing the coding gene, a co-expression engineering bacterium and an application of the co-expression engineering bacterium. Compared with a wild type enzyme, the carbonyl reductase mutant has high activity and high stereoselectivity, the enzyme activity can reach more than two times that of the wild type enzyme, the catalytic efficiency on a substrate precursor ketone is remarkably improved, the yield of a statin drug intermediate and chiral alcohol synthesized by an enzyme method of the carbonyl reductase mutant is remarkably improved, and the yield of a rosuvastatin intermediate (3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R, 3R) is remarkably improved. The highest yields of the (3R, 5R)-6-chloro-3, 5-dihydroxyhexanoic acid tert-butyl ester and the atorvastatin intermediate (3R, 5R)-6-cyano-3, 5-dihydroxyhexanoic acid tert-butyl ester reach 99% and 97.6% respectively, e.e. Is larger than 99%, and the method has high industrial application value.

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 present disclosure, among other things, provides technologies for synthesis, including reagents and methods for stereoselective synthesis. In some embodiments, the present disclosure provides compounds useful as chiral auxiliaries. In some embodiments, the present disclosure provides reagents and methods for oligonucleotide synthesis. In some embodiments, the present disclosure provides reagents and methods for chirally controlled preparation of oligonucleotides. In some embodiments, technologies of the present disclosure are particularly useful for constructing challenging internucleotidic linkages, providing high yields and stereoselectivity.

The invention discloses a dihydrodaidzein reductase mutant and application of the dihydrodaidzein reductase mutant in synthesis of (3S, 4R)-tetrahydrodaidzein, and belongs to the technical field of biology. According to the mutant, wild-type dihydrodaidzein reductase DHDR is taken as a parent, single-point mutation or combined mutation is carried out, and a DHDR mutant is obtained; the parent site of the single point mutation is one of the 120th site, the 122nd site, the 170th site, the 178th site, the 214th site, the 219th site, the 220th site or the 220th site; the combined mutation adopts two modes: one mode is that a single-point mutation DHDR mutant is taken as a parent, mutation is carried out, and a combined mutation mutant I is obtained; and 2, taking the DHDR combined mutation mutant I as a parent, and performing mutation to obtain a combined mutation mutant II. The DHDR mutant disclosed by the invention is used for synthesizing (3S, 4R)-tetrahydrodaidzein by reducing dihydrodaidzein, and asymmetric reduction with high activity and high stereoselectivity can be realized.

The invention discloses an aldehyde reductase mutant and application thereof in synthesis of a dexmethylphenidate hydrochloride intermediate, and belongs to the field of molecular biology and enzyme engineering. The aldehyde reductase mutant, polynucleotide for coding the mutant, and the recombinant expression vector can express the aldehyde reductase mutant and are used for constructing a recombinant cell or a recombinant strain for expressing the aldehyde reductase mutant. The provided aldehyde reductase mutant can catalyze 2-phenyl-2-((R)-piperidine-2)-acetaldehyde into (R)-2-phenyl-2-((R)-piperidine-2)-1-ethanol, especially improves the stereoselectivity of (R)-2-phenyl-2-((R)-piperidine-2)-1-ethanol, solves the problems of strict conditions, complex reaction and high cost in the existing synthesis method, and has a wide application prospect in the field of synthesis of (R)-2-phenyl-2-((R)-piperidine-2)-1-ethanol. Wide application prospects are realized.

The invention discloses a carbonyl reductase mutant as well as a preparation method and application thereof, and belongs to the technical field of gene engineering and enzyme engineering. According to the invention, wild-type carbonyl reductase of Stachybotrys chlorohallonata is taken as an evolution template, and a plurality of mutants with catalytic activity, thermal stability and substrate spectrum obviously superior to those of the wild-type carbonyl reductase are obtained through an enzyme engineering technology. The carbonyl reductase mutant can catalyze various prochiral carbonyl compounds to obtain chiral alcohol with high stereoselectivity (ee is greater than 99%). The carbonyl reductase mutant provides a novel, efficient and environment-friendly technical route for industrial production of chiral alcohol products, and has very high industrial application potential.

The invention discloses an enzyme catalytic synthesis method of a luliconazole chiral intermediate, which comprises the following steps: adding a recombinant escherichia coli wet cell for expressing alcohol dehydrogenase, a recombinant escherichia coli wet cell for expressing formate dehydrogenase, a buffer solution, a substrate, a cosolvent, ammonium formate and a coenzyme into a reaction container, reacting at 25-35 DEG C for 2-24 hours, extracting, separating, and carrying out rotary evaporation, thereby obtaining the luliconazole chiral intermediate. The luliconazole chiral intermediate is obtained; the substrate is 2, 2 ', 4'-trichloroacetophenone; the alcohol dehydrogenase is a mutant of AKR3. The method is mild in reaction condition, environmentally friendly, high in regioselectivity and stereoselectivity, high in conversion rate, high in chiral purity, small in enzyme dosage, low in preparation cost and suitable for industrial production.

A pharmaceutical composition comprises a GPCR agonist fusion protein in which a GPCR agonist peptide is covalently coupled to albumin via a linker in a manner that is resistant to a retro-Michael addition. Advantageously, compositions presented herein avoid decoupling of the agonist form the albumin while retaining the agonist in a steric relationship to the albumin that allows for effective binding and activation of the GPCR while also enabling gp60-mediated transcytosis and FcRn-mediated albumin recycling. These properties enable ultra-low dosages for the GPCR agonist fusion protein to give a therapeutic effect while substantially reducing or even entirely avoiding adverse effects otherwise commonly associated with unbound agonists. Such retro-Michael resistant composition is generally achieved by conformational modification of the albumin, resulting in stereoselective coupling of the linker to the albumin.

A pharmaceutical composition comprises a GPCR agonist fusion protein wherein the GPCR agonist peptide is covalently conjugated to an albumin via a linker in a manner that is resistant to a reverse Michael addition. Advantageously, the compositions presented herein avoid uncoupling of the agonist to the albumin while maintaining the agonist and the albumin in a spatial relationship that allows efficient binding and activation of the GPCR while also enabling gp60-mediated endocytosis transport and FcRn-mediated albumin recirculation. These characteristics enable ultra-low doses of GPCR agonist fusion proteins to produce therapeutic effects while significantly reducing or even completely avoiding side effects that would otherwise be typically associated with unbound agonists. Such reverse Michael resistant compositions are typically achieved by conformationally modifying an albumin, resulting in a stereoselective coupling of a linker to the albumin.

The application discloses a method for preparing a CGRP receptor antagonistic intermediate, and belongs to the field of synthesis of medical intermediates. Compound 1 is used as a raw material, reacts with hydroxylamine hydrochloride in an organic solvent to generate an intermediate 2, and then reacts in the presence of a reducing agent to obtain a remikiren intermediate 3. The application is characterized in that a large steric hindrance silicon group is used as the R substituent in the compound 1, so that high stereoselectivity is ensured in the reduction reaction. The key intermediate obtained by using the method has high chiral purity, and the operation is simple, so that the industrial production of the product in the later stage can be realized.

The invention discloses application of double enzymes in one-step preparation of L-selenium methyl selenocysteine, and belongs to the technical field of biochemical engineering. In the method, racemic N-acetyl selenium methyl selenocysteine is used, and the L-selenium methyl selenocysteine with high optical purity is prepared in a green and efficient manner through co-catalysis of stereoselective L amino acid acylase and N-acetyl amino acid racemase. The catalyst is prepared through fermentation by adopting a one-bacterium two-enzyme method. Synchronous racemization and selective hydrolysis are realized by a one-step method, the theoretical conversion rate is 100%, the actual conversion rate is 95%, and the product is e.egt; the concentration of the substrate is 8%.

The invention belongs to the technical field of organic synthesis and medicinal chemistry, and discloses a total synthesis method of rhododendron heteroterpenin Rhododaurines A-C. The total synthesis method comprises the following steps: carrying out allyl stereoselective addition on a compound 3 and a compound 5 in a toluene solvent under the conditions of catalysis of n-butyllithium and low temperature of-78 DEG C, and constructing a chiral center to obtain a compound 6; removing a methoxy methyl (MOM) protecting group from the compound 6 under an acidic condition, cyclizing, and reacting to generate rhododaurine C (7); the rhododaurine C (7) is subjected to intramolecular [4 + 2] cycloaddition and oxidation reaction in the illumination and oxygen atmosphere of 300-385 nm, and rhododaurine B (8) and an epoxy epimer epinedodaurine B (9) are obtained; and carrying out protonation and rearrangement on the epidodaurine B (9) by virtue of acid catalysis cyclic ether, so as to generate the rhododaurine A (10). The method can overcome the defects that the content of rhododendron miscellaneous terpenoids Rhododaurines A-C in nature is extremely low, and large-scale extraction is difficult.

The invention relates to a method for preparing (S)-chiral amine through asymmetric catalysis of imine reductase, and belongs to the technical field of biological pharmacy. Comprising the following steps that imine reductase catalyzes imine to be subjected to a reductive hydrogenation reaction under the conditions of coenzyme and a coenzyme regeneration system, and the (S)-chiral amine compound is generated. According to the method, imine reductase is utilized to catalyze imine asymmetric reduction to prepare the innovative drug (S)-chiral amine compound, and compared with a traditional chemical synthesis method, the method has the advantages of being high in reaction efficiency, good in stereoselectivity, mild in reaction condition, environmentally friendly and the like, and is a method meeting the green production requirement.

The invention belongs to the technical field of asymmetric catalysis, and discloses a chiral phosphine nitrogen-nitrogen tridentate ligand based on a ferrocene skeleton as well as a preparation method and application of the chiral phosphine nitrogen-nitrogen tridentate ligand. The structural formula of the ligand is shown as (I) or an enantiomer (II) of the ligand. The preparation method comprises the steps of acylation reaction, oxazoline ring construction, aldehyde group introduction, reductive amination and the like, and application in asymmetric hydrogenation reaction, such as asymmetric hydrogenation of aryl ketone. The ligand has a plurality of chiral factors, simultaneously has a potential hydrogen bond donor, forms secondary interaction with a substrate in an asymmetric catalytic reaction, can further stabilize a reaction transition state, is an efficient novel chiral P, N, N-ligand, has the advantages of high reaction activity, good stereoselectivity, wide substrate range and the like, and has a wide application prospect. # imgabs0 #

The invention provides immobilized ketoreductase, an immobilization method and application of the immobilized ketoreductase in preparation of a key intermediate of bentonatate, the amino acid sequence of the ketoreductase is as shown in SEQ ID NO.2, preferably a ketoreductase mutant is as shown in SEQ ID NO.16, and the immobilized ketoreductase comprises resin and corresponding ketoreductase. The ketoreductase mutant and the immobilized ketoreductase prepared from the ketoreductase mutant are very high in enzyme activity during ketoreduction reaction, and when the ketoreductase mutant is used for preparing a key intermediate of bentonamate, a solvent is pure water, so that the ketoreductase mutant and the immobilized ketoreductase prepared from the ketoreductase mutant are environment-friendly. The immobilized transaminase is high in conversion rate, good in stereoselectivity, good in stability, reusable and simpler to operate, so that the production cost is reduced, and industrial production is facilitated.

The invention belongs to the technical field of synthesis of organic compounds, and particularly relates to a chiral synthesis method of spiro [cyclopenta [c] carbazole-1, 1 '-isoindoline]-3'-ketone compounds. The compound synthesized by the invention has an isoindolinone skeleton and a carbazole active fragment at the same time. The carbazole compound and the isoindolinone compound are successfully combined in a cyclization mode for the first time, and the chiral compound can be prepared. The invention provides a synthesis method of a chiral spiro [cyclopenta [c] carbazole-1, 1 '-isoindoline]-3'-ketone compound, which has the advantages of mild reaction conditions, green and environment-friendly catalyst, easiness in operation, few synthesis steps, easiness in purification and separation of products, capability of obtaining higher yield and high stereoselectivity, easiness in amplification of reaction, high yield, high yield and high yield, and is suitable for large-scale production of the chiral spiro [cyclopenta [c] carbazole-1, 1 '-isoindoline]-3'-ketone compound. And the method has a wide application prospect.

To provide a novel stereoselective method for producing a cyclic dinucleotide derivative that can be used for an antibody-immunostimulator conjugate, and a production intermediate thereof, and to provide methods for producing a cyclic dinucleotide-linker and antibody-immunostimulator conjugate using the production method.SOLUTION: The present invention relates to a method that makes it possible to produce, in large amounts and with high yield, a cyclic dinucleotide derivative having a desired steric configuration, by using an optically active phosphitylation agent twice, during coupling and cyclization.SELECTED DRAWING: None

The invention relates to a carbonyl reductase mutant based on cosubstrate catalytic ability optimization and application thereof, the mutant is constructed on the basis of a carbonyl reductase BaSDRX amino acid sequence as shown in SEQ ID No.2, the nucleotide sequence of the carbonyl reductase is as shown in SEQ ID No.1 in a sequence table, and the nucleotide sequence of the carbonyl reductase is as shown in SEQ ID No.2 in the sequence table. The mutant is single-point mutation or multi-point combined mutation on the basis of an amino acid sequence of carbonyl reductase BaSDRX as shown in SEQ ID No.2: threonine Thr133 at the 133 site, arginine Arg137 at the 137 site and glutamic acid Glu142 at the 142 site; when the mutant enzyme or a recombinant cell containing the mutant enzyme is applied to synthesis of (S)-1-(2, 6-difluorophenyl) ethanol through asymmetric reduction of 2, 6-difluoroacetophenone, high catalytic activity and high stereoselectivity are achieved, and high-optical-purity (S)-1-(2, 6-difluorophenyl) ethanol (eet; therefore, the method has a good industrial application and development prospect.

The invention relates to a synthesis method of milobalin or acid salt thereof, and belongs to the technical field of medicine synthesis. In order to solve the problem that chiral resolution needs to be adopted in the prior art, the invention provides a synthesis method of miloabalin or an acid salt thereof, which comprises the following steps: under the action of an organic strong base, carrying out Michael addition reaction on a compound shown as a formula III and acetonitrile in a non-polar organic solvent at a low temperature of-50 DEG C or below to obtain an intermediate product; under the alkaline condition, the intermediate product is subjected to an oxidation reaction and a hydrolysis reaction under the action of an oxidizing agent, after the reaction is finished, an intermediate product is obtained through acidification, and the intermediate product is subjected to a high-temperature decarboxylation reaction in an aprotic solvent to obtain a compound of a formula VI; and carrying out Hofmann degradation reaction on the compound of formula VI to obtain the compound of formula I miloabalin or miloabalin acid salt. The method has the advantages of high stereoselectivity, no need of chiral resolution operation, avoidance of the problem of large material loss caused by resolution, high chiral purity and high yield.

The invention discloses a method for selectively depolymerizing polylactic acid into chiral lactide, and belongs to the technical field of polylactic acid depolymerization. According to the invention, the problems that the existing conventional polylactic acid depolymerization means generally lacks stereoselectivity and a depolymerization product with high optical purity is difficult to obtain are solved. In the presence of a solvent and under the heating condition, polylactic acid matched with chirality is depolymerized into lactide with high optical purity under the catalysis of a chiral depolymerization catalyst, and polylactic acid not matched with chirality is not depolymerized, so that the selective depolymerization process of polylactic acid is realized. Different from the traditional asymmetric catalysis field, the method focuses on recognition of small molecules, realizes closed-loop depolymerization of polylactic acid to chiral monomers, realizes selective recognition and depolymerization of chiral polymers, and broadens the research range of asymmetric catalysis.

The invention relates to the technical field of biology, in particular to a carbonyl reductase mutant, a gene, an engineering bacterium and application. The amino acid sequence of the carbonyl reductase mutant is shown as SEQ ID NO: 1, the catalytic efficiency of asymmetric reductive hydrogenation of prochiral ketone compounds is improved, the stereoselectivity of chiral hydroxyl compounds formed through catalysis is improved, and the synthesis efficiency of (S)-1-(5-fluoro-2-iodophenyl) ethanol is improved.

The invention discloses an oxidoreductase mutant and application thereof, and belongs to the field of enzyme engineering. The invention provides a recombinant oxidoreductase HbKR mutant which is used for catalytically producing (R)-decalactone. The oxidoreductase HbKR mutant disclosed by the invention realizes soluble expression in Escherichia coli BL21 (DE3), and the oxidation activity of the oxidoreductase HbKR mutant is improved by 4.61 times compared with that of a parent enzyme. An oxidoreductase HbKR mutant and lactone hydrolase Rpa3624 derived from Rhodopseudomonas palustris are combined for use, reaction is performed for 0.5 h under the conditions that the temperature is 35 DEG C and the pH value is 10.5, compared with a wild type, the yield of (R)-delta-decalactone is increased to 48.2% from 42.3%, and the stereoselectivity is increased to 98.7% from 94.2%.

The invention provides a method for synthesizing N-methylated polypeptide, which comprises the following steps: dissolving a carboxylic acid compound, N-methyl amino-acid ester hydrochloride or N-methyl amino-acid ester, an alkaline substance and pivaloic anhydride in an organic solvent, reacting at 15-80 DEG C for 3-10 hours, and post-treating the obtained reaction liquid to obtain an N-methyl dipeptide compound, the method comprises the following steps: removing a protecting group in an N-methyl dipeptide compound to obtain a free-state N-methyl dipeptide compound, and condensing the free-state N-methyl dipeptide compound and amino acid or N-methyl amino-acid ester protected by amino to obtain the N-methylated polypeptide, the mixed anhydride intermediate is formed in situ, and the reaction is completed in one step; according to the method, trimethylacetic anhydride is used as a condensing agent, the structure is simple, the reaction is safe, cheap and non-toxic, a large amount of nitrogen-containing byproducts are not generated in the reaction, and purification is convenient; the method is high in stereoselectivity, and racemization is avoided; according to the method, environment-friendly solvents such as ethyl acetate can be used, the reaction condition is mild, and the reaction time is short.

The application discloses a carbonyl reductase mutant and application thereof in preparation of (R)-8-chloro-6-hydroxyoctanoate compounds. The application relates to a mutant of carbonyl reductase with improved catalytic performance and stereoselectivity after rational molecular modification, a recombinant expression plasmid containing the mutant gene of the enzyme, a recombinant expression transformant, a preparation method of the recombinant enzyme, and application of the recombinant enzyme or the recombinant cell as a catalyst in asymmetric reduction of 8-chloro-6-carbonyloctanoate compounds (I) to prepare (R)-8-chloro-6-hydroxyoctanoate compounds (II), in particular, in an intermediate (R)-8-chloro-6-hydroxyoctanoate methyl ester of thioctic acid. The disclosed carbonyl reductase SsCR mutant has the advantages of high reaction efficiency and high catalytic activity, and exhibits the advantages of high substrate concentration, low catalyst consumption and short reaction time in the process of catalytic preparation of the compound (II).

The present invention discloses a transaminase and its use in preparing optically pure chiral amines. The amino acid sequence of the transaminase of the present invention has one or more mutations at positions 65, 318, 399, 419, 426, 465, and 466 compared to SEQ ID NO: 1, wherein the mutation is an addition, deletion, or substitution of an amino acid residue. The present invention also discloses a nucleic acid molecule encoding the transaminase, a nucleic acid construct comprising the nucleic acid molecule, a recombinant vector, and a host cell. The present invention also discloses an enzyme preparation comprising the transaminase of the present invention. The present invention also discloses a method for preparing chiral amines using the transaminase. In the industrial production process of optically pure chiral amines, the enzyme provided by the present invention has advantages such as substrate specificity, enantiomeric selectivity, and high conversion rate. The method for preparing optically pure chiral amines provided by the present invention has high reaction efficiency, stereoselectivity, and yield.

The invention discloses a halohydrin dehalogenase mutant and application of the halohydrin dehalogenase mutant in preparation of (R)-4-phenyl-2-oxazolidinone, and belongs to the technical field of enzyme engineering and chemical engineering, amino acid residues of wild type halohydrin dehalogenase are mutated, and the halohydrin dehalogenase mutant with high enzyme activity and high stereoselectivity is screened out. The halohydrin dehalogenase mutant provided by the invention has high enzyme activity and high stereoselectivity in the application of catalyzing (S)-2-phenyl ethylene oxide to be converted into (R)-4-phenyl-2-oxazolidinone, the catalytic efficiency of the mutant T126L / F170M can reach 1.7 times of that of wild halohydrin dehalogenase, and the halohydrin dehalogenase mutant has extremely high industrial application value.

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 present application discloses a chiral dienyl-substituted carboxylate compound and an asymmetric preparation method thereof, wherein the chiral dienyl-substituted carboxylate compound is an optically active compound with the structure shown in the following formula I, including stereoisomers with the same chemical formula: in the formula, * represents a chiral carbon atom; the substituent R is selected from hydrogen, C 1‑10 alkyl or aryl; Ar is an aromatic ring or a substituted aromatic ring; R 1 , R 2 , R 3 , R 4 , R 5 are independently selected from hydrogen, halogen, C 1‑10 alkyl or alkoxy; R 6 is independently selected from C 1‑10 alkyl or aryl; R 7 is independently selected from C 1‑10 alkyl or aryl. The present application applies a chiral isothiourea catalytic electrochemical synthesis system to precisely and quickly synthesize the chiral dienyl-substituted carboxylate compound in a high yield and high stereoselectivity and in a gram-scale. The preparation method of the present application is simple, and has biomedical practicability and industrial application prospect.

The invention provides an R-type transaminase mutant and application thereof, and compared with an amino acid sequence of wild-type transaminase as shown in SEQ ID NO.1, the mutant has mutation of a 62nd site, a 152nd site and a 155th site. The mutant has high catalytic activity and excellent stereoselectivity on a heterocyclic ketone substrate, and can be used for efficiently preparing R-configuration heterocyclic chiral amine.

The invention discloses a synthetic intermediate of entecavir, a preparation method and application. The intermediate can be used for asymmetrically synthesizing optically pure (+)-entecavir. By utilizing the intermediate, asymmetric synthesis of entecavir can be efficiently and accurately realized, which is of great significance for improving the stereoselectivity of the product and improving the synthesis efficiency. The synthesis method of the entecavir and the intermediate thereof has the advantages of cheap and easily available reagents, simple synthesis steps, high reaction yield and the like, and is simple in reaction process, convenient to operate, green, environment-friendly and suitable for industrial large-scale production.