Method for producing tyrosol and hydroxytyrosol through heterologous metabolic pathways

A technology for hydroxytyrosol and tyrosol, applied in the field of heterologous metabolic pathway production of tyrosol and hydroxytyrosol, can solve the problems of long cycle, large environmental pollution, lack of production methods and the like

Active Publication Date: 2018-01-16
BEIJING UNIV OF CHEM TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the lack of effective production methods, hydroxytyrosol cannot be produced on a large scale
At present, hydroxytyrosol is mainly derived from olive leaf extract, or extracted from olive oil or olive oil factory wastewater, but these processes have low recovery r

Method used

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  • Method for producing tyrosol and hydroxytyrosol through heterologous metabolic pathways
  • Method for producing tyrosol and hydroxytyrosol through heterologous metabolic pathways
  • Method for producing tyrosol and hydroxytyrosol through heterologous metabolic pathways

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Experimental program
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Effect test

specific Embodiment 1

[0030] Production of tyrosol and hydroxytyrosol by adding tyrosine in vitro

[0031] The tyrosine directly added to the medium during the fermentation process is screened from bacteria, fungi or protein engineered aminotransferase (AT), ketoacid decarboxylase (KDC), alcohol dehydrogenase (ADH) to catalyze the formation of tyrosine Alcohol, and on this basis, 4-hydroxyphenylacetic acid 3-monooxygenase (HpaBC) is introduced to catalyze the formation of hydroxytyrosol. Aminotransferase (AT), ketoacid decarboxylase (KDC), alcohol dehydrogenase (ADH), 4-hydroxyphenylacetic acid 3-monooxygenase (HpaBC) gene fragments were obtained by PCR, followed by restriction endonuclease Enzyme digestion of gene fragments and plasmid vectors, gel recovery or column recovery of the digested fragments, and then aminotransferase (AT), ketoacid decarboxylase (KDC), alcohol dehydrogenase (ADH) target gene Insert into plasmid pZE12-luc (high copy) to obtain pZE-ARO10-ADH6 recombinant plasmid; insert ...

specific Embodiment 2

[0035] Construction of Metabolic Pathways for Microbial Heterologous Synthesis of Tyrosol and Hydroxytyrosol

[0036] Starting from glucose and glycerol, tyrosine is synthesized through the shikimate pathway, and tyrosine is screened from bacteria, fungi or protein engineered aminotransferase (AT), ketoacid decarboxylase (KDC), alcohol dehydrogenation The enzyme (ADH) catalyzes the generation of tyrosol, and on the basis of tyrosol, 4-hydroxyphenylacetic acid 3-monooxygenase (HpaBC) is introduced to catalyze the reaction of tyrosol to hydroxytyrosol. Aminotransferase (AT), ketoacid decarboxylase (KDC), alcohol dehydrogenase (ADH), 4-hydroxyphenylacetic acid 3-monooxygenase (HpaBC) and TyrA that enhances the shikimate pathway were obtained by PCR fbr , PpsA, TktA and AroG fbr Enzyme gene fragments, then use restriction endonucleases to digest the gene fragments and plasmid vectors, perform gel recovery or column recovery on the digested fragments, and then insert the target ge...

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Abstract

The invention discloses a method for producing tyrosol and hydroxytyrosol through heterologous metabolic pathways, wherein metabolic pathways for producing hydroxytyrosol are introduced into hosts forproducing the hydroxytyrosol; more specifically, the method comprises the following steps: producing tyrosol by efficiently expressing aminotransferase, ketoacid decarboxylase and alcohol dehydrogenase in the hosts, then introducing 4-hydroxyphenylacetate hydroxylase to produce the hydroxytyrosol. Genes coded by the enzymes are constructed on an expression plasmid or integrated to a genome. Through the method, production hosts capable of producing the hydroxytyrosol by using glucose, glycerinum and tyrosine can be prepared by introducing the genes coded by the enzymes to the hosts; meanwhile,the invention also discloses a biphasic cultivation method, so that the production of the hydroxytyrosol is more efficient.

Description

technical field [0001] The present invention relates to the field of biotechnology, more precisely, the present invention relates to heterologous biosynthesis of tyrosol, the metabolic pathway of hydroxytyrosol and the method of biphasic culture, by enhancing the expression of upstream shikimate pathway enzymes, selecting the source from bacteria, The exogenous enzyme modified by fungi or protein engineering uses original or modified bacteria, fungi, plant cells or animal cells as host cells, and finally improves the production of tyrosol and hydroxytyrosol. Background technique [0002] Tyrosol is one of the main active compounds in Rhodiola, which has significant effects in anti-hypoxia, anti-fatigue, anti-radiation, anti-cancer, anti-aging, and regulation of body immunity. It is mainly used as a compound in the synthesis of medicine and spices. Intermediates, such as synthetic cardiovascular drugs Metoprolol, Betasolol, etc. Hydroxytyrosol (HT) is a natural phenolic comp...

Claims

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Application Information

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IPC IPC(8): C12P7/22
Inventor 孙新晓袁其朋李向来陈振娅
Owner BEIJING UNIV OF CHEM TECH
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