CN-122012554-A - Atractylodes lancea alpha-eucalyptol synthase gene AlTPS alpha, its coding product and application

Abstract

The invention discloses a atractylis lancea alpha-eucalyptol synthase gene AlTPS alpha, a coding product and application thereof. The nucleotide sequence of the atractylis lancea alpha-eucalyptol synthase is shown as SEQ NO.1, and the coding amino acid sequence is shown as SEQ NO. 2. The method has the activity of generating the terpenoid alpha-eucalyptol by taking farnesyl pyrophosphate FPP as a substrate. The alpha-eucalyptol is a volatile pharmacologic active ingredient of various plants, and has the effects of tranquilizing and allaying excitement and the like. The alpha-eucalyptol synthase coding gene provided by the invention can be used for efficiently producing alpha-eucalyptol by a microbial engineering technology.

Inventors

• HUANG XIAO
• LIU YUEZHEN
• YU KUN
• ZHANG MENGYUAN
• LI TINGTING
• CHEN LEI

Assignees

• 湖北中医药大学

Dates

Publication Date

20260512

Application Date

20260204

Claims (10)

1. 1. The atractylis lancea alpha-eucalyptol synthase encoding gene is characterized in that the encoding nucleotide sequence is shown as SEQ NO. 1.
2. 2. The amino acid sequence of atractylis lancea α -eucalyptol synthase according to claim 1.
3. 3. The amino acid sequence according to claim 2, wherein the amino acid sequence is shown in SEQ No. 2.
4. 4. A recombinant vector comprising the coding gene or the translated amino acid of claim 1,2 or 3.
5. 5. The recombinant vector according to claim 4, wherein the vector is pET28a.
6. 6. A recombinant strain comprising the coding gene of claim 1 or 2 or 3 and translated amino acids.
7. 7. A host cell comprising the coding gene of claim 1 or 2 or 3 and translated amino acids.
8. 8. A method for expressing alpha-eucalyptol synthase and producing alpha-eucalyptol, which is characterized in that the method comprises transforming a strain suitable for the expression thereof with the recombinant vector of claim 4 or 5 to obtain the recombinant strain or host cell of claim 6 or 7, culturing the recombinant strain or host cell, inducing the expression of the alpha-eucalyptol synthase of claim 2 or 3, synthesizing FPP substrate by the recombinant strain at the same time, synthesizing alpha-eucalyptol by the alpha-eucalyptol synthase at the same time using FPP, and recovering the alpha-eucalyptol product thereof after the end of the culture.
9. 9. Use of the coding gene according to claim 1, or the coding amino acid sequence according to claim 2 or 3, or the recombinant vector according to claim 4 or 5, or the recombinant strain according to claim 6, or the host cell according to claim 7, for the preparation of α -eucalyptol.
10. 10. The use according to claim 9, wherein the terpenoid is α -eucalyptol.

Description

Atractylodes lancea alpha-eucalyptol synthase gene AlTPS alpha, its coding product and application Technical Field The invention relates to the technical field of plant genetic engineering and natural product biology, in particular to a atractylis lancea alpha-eucalyptol synthase gene AlTPS alpha, and a coding product and application thereof. Background The terpenoid is one of the secondary metabolites with the most variety and most structural diversity in natural products, and has wide application value in the fields of medicine, spice, food, agriculture and the like. Terpene synthases (TERPENE SYNTHASE, TPS) are key enzymes in the biosynthesis pathway of terpenes, and can catalyze precursor substances such as geranylgeranyl pyrophosphate (GGPP) or farnesyl pyrophosphate (FPP) to generate basic terpene skeletons with different structures, so that various active terpenes are formed through the action of modified enzymes. At present, terpene synthase genes of various plants are cloned and functionally identified, but the catalytic specificity, efficiency and expression regulation mechanism of the terpene synthase genes still have great differences, so that the terpene synthase genes with specific catalytic functions are mined and functionally represented, and the terpene synthase genes have important significance for analyzing the biosynthesis pathway of terpenes and realizing efficient and directional production of target active ingredients. Alpha-eucalyptol is one of the main active ingredients of rhizoma atractylodis (Atractylodes lancea (thunder.) DC.) and belongs to sesquiterpenoids, and has various pharmacological activities such as anti-inflammatory, antibacterial and neuroprotection. At present, the production of alpha-eucalyptol mainly depends on plant extraction, but is limited by factors such as long plant growth period, low content, complex extraction process and the like, and the market demand is difficult to meet. The cell factory for efficiently producing the alpha-eucalyptol is constructed by utilizing a synthetic biological strategy, is an important way for realizing sustainable mass production, and cloning and obtaining the efficient and specific alpha-eucalyptol synthase gene are core preconditions for realizing the strategy. However, cloning and functional verification of an alpha-eucalyptol synthase gene in rhizoma atractylodis are not reported at present, and catalytic properties and application potential of the coded enzyme protein are still to be studied intensively. Disclosure of Invention In view of the defects of the prior art, the invention provides a atractylis lancea alpha-eucalyptol synthase gene AlTPS alpha, and a coding product and application thereof. The coding gene of the atractylis lancea alpha-eucalyptol synthase provided by the invention can realize sustainable large-scale production of alpha-eucalyptol. In one aspect of the invention, a atractylis lancea alpha-eucalyptol synthase encoding gene is provided, and the encoding nucleotide sequence of the gene is shown as SEQ NO. 1. In another aspect of the invention, a atractylis lancea alpha-eucalyptol synthase is provided, and the amino acid sequence of the atractylis lancea alpha-eucalyptol synthase is shown as SEQ NO. 2. In another aspect of the present invention, there is provided a recombinant expression vector comprising the foregoing coding gene. Preferably, the expression vector is pET-28a. In another aspect of the present invention, there is provided a recombinant strain comprising the foregoing coding gene. In another aspect of the invention, there is provided a host cell comprising the foregoing encoding gene. In another aspect of the present invention, there is provided a method for expressing atractylis lancea α -eucalyptol synthase, comprising transforming a strain suitable for its expression with the aforementioned recombinant vector to obtain a recombinant strain, culturing the recombinant strain, inducing the expression of atractylis lancea α -eucalyptol synthase, synthesizing an FPP substrate by the recombinant strain simultaneously, synthesizing α -eucalyptol by the α -eucalyptol synthase simultaneously using FPP, and recovering the synthesized α -eucalyptol product after the culturing is completed. In another aspect of the invention, the use of the aforementioned coding gene, or recombinant vector, or recombinant strain, or host cell, in the preparation of terpenoids is provided. Preferably, the terpenoid product is α -eucalyptol. The invention has the beneficial effects that: A terpene synthase gene in rhizoma atractylodis lanceae is obtained, and the coded protein has the activity of generating a terpenoid alpha-eucalyptol by taking FPP as a substrate. The invention also provides a method for directly preparing the terpenoid in the recombinant strain. The gene and the preparation method provided by the invention can realize sustainable large-scale production of alpha-eucalyptol. Drawings The invention