Novosphingobium short-chain alcohol dehydrogenase mutant and application thereof

A new sphingolipid and short-chain alcohol technology, applied in the direction of enzymes, oxidoreductases, enzymes, etc., can solve the problems of harsh reaction conditions and low chiral ee value

Active Publication Date: 2018-12-28
ZHEJIANG OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] Aiming at the problem that (R)-β-hydroxymyristate is prepared by chiral catalysts, the chiral ee value is relatively low, and the reaction conditions are relatively harsh. The purpose of the present invention is to provide a short-chain Alcoho

Method used

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  • Novosphingobium short-chain alcohol dehydrogenase mutant and application thereof
  • Novosphingobium short-chain alcohol dehydrogenase mutant and application thereof
  • Novosphingobium short-chain alcohol dehydrogenase mutant and application thereof

Examples

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Effect test

Embodiment 1

[0042] A short-chain alcohol dehydrogenase mutant from Neosphingomum marinum, which uses the amino acid sequence of NA-ADH of Neosphingomum marinum short-chain alcohol dehydrogenase as a template, and transforms the first amino acid sequence of NA-ADH according to site-directed mutagenesis technology. NA-ADH-M1 was constructed by mutating glycine at position 145 to alanine.

Embodiment 2

[0044] A short-chain alcohol dehydrogenase mutant from Neosphingomum marinum, which uses the amino acid sequence of NA-ADH of Neosphingomum marinum short-chain alcohol dehydrogenase as a template, and transforms the first amino acid sequence of NA-ADH according to site-directed mutagenesis technology. The 199-position isoleucine was mutated to leucine to construct NA-ADH-M2.

Embodiment 3

[0046] A short-chain alcohol dehydrogenase mutant from Neosphingomum marinum, which uses the amino acid sequence of NA-ADH of Neosphingomum marinum short-chain alcohol dehydrogenase as a template, and transforms the first amino acid sequence of NA-ADH according to site-directed mutagenesis technology. NA-ADH-M3 obtained by mutating glycine at position 145 to alanine and isoleucine at position 199 to leucine.

[0047] Application of the N. marine short-chain alcohol dehydrogenase mutant obtained from the above-mentioned N. marine N. sphingolis short-chain alcohol dehydrogenase NA-ADH in the preparation of (R)-beta-hydroxy myristate, wherein The structural formula of (R)-β-hydroxytetradecanoate is:

[0048]

[0049] R in the molecular structural formula represents a saturated alkyl group with 1 to 8 carbon atoms, and (R) represents chirality. Taking the preparation of (R)-β-hydroxytetradecanoic acid methyl ester and (R)-β-hydroxytetradecanoic acid ethyl ester as an example, ...

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Abstract

The invention relates to the technical field of dehydrogenase mutants, in particular to a novosphingobium short-chain alcohol dehydrogenase mutant and an application thereof. The novosphingobium short-chain alcohol dehydrogenase mutant adopts the amino acid sequence of novosphingobium short-chain alcohol dehydrogenase NA-ADH as a template. The short-chain alcohol dehydrogenase mutant of a marine origin is obtained by site-directed mutagenesis. The mutuant is transferred into an expression strain BL21 (DE3) to obtain thalli containing the recombinant short-chain alcohol dehydrogenase after recombination by an expression vector, and then beta-hydroxytetradecanoate is used as a substrate to prepare (R)-beta-hydroxytetradecanoate by bio-enzyme catalysis, the chiral ee value of the obtained (R)-beta-hydroxytetradecanoate can reach above 99, and the substrate conversion rate is above 95%. The preparation conditions are mild, and the process way is simple.

Description

technical field [0001] The invention relates to the technical field of dehydrogenase mutants, in particular to a short-chain alcohol dehydrogenase mutant of Neosphingomum marine and its application. Background technique [0002] Orlistat is a potent and long-acting specific gastrointestinal lipase inhibitor. By directly blocking the body's absorption of fat in food, the intake of heat energy and fat is less than the consumed fat, thereby reducing the body's Fat achieves the purpose of weight loss, and the active ingredients of orlistat do not enter the blood circulation and do not act on the central nervous system, so there are fewer side effects, and there will be no dizziness, palpitation, insomnia, dry mouth, etc. It is currently the only OTC in the world Diet Pills and Top Selling Weight Loss Products. The structural formula of orlistat is: [0003] [0004] It can be seen from the analysis that (R)-β-hydroxy myristate is an important raw material for the synthesis ...

Claims

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Application Information

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IPC IPC(8): C12N9/04C12P7/64
CPCC12N9/0006C12P7/6436C12Y101/01
Inventor 唐云平周亚峰丁国芳余方苗杨最素黄芳芳陈艳
Owner ZHEJIANG OCEAN UNIV
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