33 results about "Amine dehydrogenase" patented technology

Non-naturally occurring amine dehydrogenases (AmDH) and methods of use thereof the produce chiral amines are disclosed. The AmDH are variants of amino acid dehydrogenases. AmDH based on phenylalanine, leucine, and valine scaffolds are provided. The AmDH typically have one, two, three, four, or more amino acid alterations relative to the scaffold. The alterations to the scaffold result in an enzyme that accepts the analogous ketone, such as methyl isobutyl ketone (MIBK), instead of the wild-type α-keto acid. Chimeric AmDH are also disclosed. The chimeras are fusion proteins that include a substrate binding domain from a first AmDH and a cofactor binding domain from a second AmDH. In a preferred embodiment, one of the domains is from a PheDH-based AmDH and one of the domains is from a LeuDH-based AmDH.

The invention relates to a method and a kit for stably detecting sialic acid by an enzyme method. The method comprises the following steps of: generating N-acetylneuraminic acid from bound sialic acid under the catalysis of neuraminic acid aldolase; transforming the N-acetylneuraminic acid into N-acetylmannosamine under the catalysis of neuraminidase; reacting the product with mannosamine dehydrogenase and nicotinamide adenine dinucleotide (NAD) to generate reduced nicotinamide adenine dinucleotide (NADH); and measuring the sialic acid content by detecting the amount of the generated NADH. The invention also relates to the kit prepared according to the method. The method and the kit are conveniently and quickly used, the sialic acid is not required to be redissolved by dissolving liquid, is stable for a long time and has small inter-bottle variation, the method and the kit are suitable for various biochemical analyzers, secondary pollution is avoided, and a production process method is simplified.

The invention provides a method for preparing chiral amine through asymmetric reduction under the catalysis of a marine strain, and relates to chiral amine. The method comprises the following steps: inoculating a strain into a 216L culture medium, culturing, then centrifuging the obtained fermentation liquid to obtain cells, and redissolving and washing with a buffer solution, thus preparing cell sap; and adding ketone used as a substrate into the obtained cell sap, then adding an amino donor and a cosubstrate for cyclic regeneration of coenzyme, and performing asymmetric reduction and amination under the catalysis of the cells, thus obtaining the chiral amine product. The new strain which contains NADH (nicotinamide adenine dinucleotide)-dependent amine dehydrogenase and is different from strains reported before is obtained through the screening of marine microorganisms, and the reaction system and reaction conditions of preparing chiral amine through the asymmetric reduction and amination of ketone under the catalysis of full cells of the strain are optimized. The optical purity of the obtained chiral amine product is up to 90% or above, and the yield is up to 70% or above. The method provided by the invention is convenient to operate, has the advantages of high optical purity of the product, high yield and the like, is simple in equipment and has better industrial application prospects in the field of preparing chiral amine under biological catalysis.

The invention discloses an amine dehydrogenase mutant and its application in the synthesis of chiral aminoalcohol compounds. The amine dehydrogenase mutant disclosed in the present invention is the 40th, 61st, 68th, 69th, 111th, 113th, 114th, 116th to the amine dehydrogenase (SpAmDH) derived from psychrophilic bacillus sarcina (Sporosarcina psychrophila) , 134, 150, 187, 239, 288, 291, 292, 295, 297 and / or 294 positions are mutated. The amine dehydrogenase mutant of the present invention can catalyze the synthesis of hydroxyketone substrates into chiral aminoalcohol compounds represented by 1-hydroxy-2-butanone and 4-hydroxy-2-butanone, and is easy to operate and has optical purity of the product High advantages, the stereoselectivity can be as high as 99%, and the biocatalytic preparation of chiral aminoalcohol compounds has good industrial application prospects.

The invention discloses a synthesis method of a chiral aminoalcohol compound. The application of the biological material comprising amine dehydrogenase provided by the present invention in at least one of the following A1-A6); A1) catalyzing 1-hydroxyl-2-butanone to generate (S)-2-amino-1-butanol ; A2) catalyzing 4-hydroxyl-2-butanone to generate (R)-3-amino-1-butanol; A3) synthesis or preparation of chiral amino alcohol compounds; A4) synthesis or preparation of 2-amino-1-butanol and 3‑amino‑1‑butanol; A5) synthesis or preparation of (S)‑2‑amino‑1‑butanol and (R)‑3‑amino‑1‑butanol; A6) catalysis of hydroxyketone substrates to generate aminoalcohol compounds The experiment proves that the method provided by the present invention has the advantages of mild reaction conditions, good stereoselectivity, high yield and simple operation.

The invention discloses a transformation strain and a method for catalyzing the synthesis of miglitol intermediates. The transformation strain is obtained from the starting strain ZJB16009 by transforming the recombinant expression plasmid pBBR-sldAB, and then screening, named Gluconobacter oxidans ( Gluconobacter oxydans), strain number pBBR‑sldAB, preservation number CCTCC No.M 2019033, its N‑hydroxyethylglucosamine dehydrogenase activity is greatly improved compared with the starting strain ZJB16009, and the space-time yield is about that of the starting strain ZJB16009 2 times, it shows that the Gluconobacter oxydans pBBR-sldAB of the present invention has higher potential application value in the catalytic synthesis of the key intermediate 6NSL of miglitol.

The invention discloses a marine bacterial strain and a method for preparing chiral amine from catalyzing of amine dehydrogenase of the marine bacterial strain and belongs to the technical field of asymmetric synthesis chiral compounds obtained through a biological method. Psedomonas balearica from the sea is obtained through screening, and under the condition that ketone serving as a substrate, an amino donor and an auxiliary substrate are added, the chiral amine is prepared by the utilization of asymmetric reductive amination of microorganism and crude enzyme catalyzed ketone of the bacterial strain. A reaction system and reaction conditions of asymmetric reductive amination of microorganism catalyzing and crude enzyme catalyzing of the strain are determined. The method has good industrial application prospects in the field of preparation of chiral amine through biological catalysis and has important meaning for research on development of marine novel biocatalyst and a green synthesis method for chiral amine in the future.

The invention relates to polyethylene imine-titanium oxide embedded with amine dehydrogenase and a preparation method of the polyethylene imine-titanium oxide, and belongs to the technical field of immobilized enzymes. A polyethylene imine (PEI)-titanium oxide immobilized particle embedded with the amine dehydrogenase has a grain size of 100-600 nm, wherein polyethylene imine has catalyzing and templating double effects in the immobilization process, namely the PEI promotes the formation of titanium oxide through an electrostatic and hydrogen bonding effect between an amino group in a molecule and a titanium precursor molecule; through a polyethylene imine structure, a template can also be induced, and the formation of a titanium oxide particle are regulated and controlled. The preparation method of the polyethylene imine-titanium oxide comprises the following steps: configuring the PEI (straight chain and branched chain) as well as mixed liquid containing the amine dehydrogenase; configuring a precursor II-dihydroxybis (2-hydroxypropanoato) titante diammonium (Ti-BALDH) solution of titanium; dropwise adding the precursor solution of the titanium into the PEI and the mixed liquid containing the amine dehydrogenase, stirring and generating the PEI-titanium oxide immobilized particle embedded with the amine dehydrogenase. The polyethylene imine-titanium oxide and the preparation method thereof have the advantages that the preparation conditions are mild, the process is simple and is easy to implement, the obtained immobilized particle embedded with the amine dehydrogenase has strong immobilization capacity and good reuse stability, and the temperature stability of the amine dehydrogenase can be obviously improved.

The invention discloses a method for preparing chiral amine from multi-enzyme coupled systems and belongs to the technical field of biological catalysis asymmetric conversion. An amine dehydrogenase and glycerol dehydrogenase coupled system, an amine dehydrogenase and formate dehydrogenase coupled system as well as an amine dehydrogenase, glycerol dehydrogenase and formate dehydrogenase coupled system are constructed, and asymmetric preparation of the chiral amine and regeneration of coenzyme nicotinamide adenine dinucleotide are realized. The multi-enzyme coupled systems and the method for preparing the chiral amine from the multi-enzyme coupled systems have advantages that the product conversion rate is high, the reaction condition is mild, operation is simple, the coenzyme can be regenerated and the like, the cost is low, the enzyme utilization rate is high, and the method is suitable for industrial production.

The invention belongs to the technical field of biology, in particular to an amine dehydrogenase mutant with improved thermal stability, a construction method and an application of genetic engineeringbacteria. The amine dehydrogenase mutant comprises 4 single-site mutants and 11 combined mutants, and compared with wild-type amine dehydrogenase, the single-site mutants and the combined mutants have longer half-life period at 42 DEG C; in particular, the combined mutants show a superposition effect of thermal stability of the single-site mutants, and the half-life period is about 5 times that of the wild-type amine dehydrogenase. The amine dehydrogenase mutant obtained by the construction method has better thermal stability, shows excellent stereoselectivity, regioselectivity and catalyticactivity when being catalyzed for synthesis of chiral amine at higher temperature, and has a better application prospect.