Application of UGT71A3 as catalyst in preparation of triterpenoid saponins

Abstract

The application provides application of UGT71A3 as a catalyst in preparation of triterpene saponins. Research finds that UGT71A3 as a catalyst can be used for preparation of triterpene saponins, and has a high yield, and provides a new method for preparation of rare triterpene saponins.

Description

Technical Field

[0001] This invention relates to the field of natural product chemistry, and in particular to the application of UGT71A3 as a catalyst in the preparation of triterpenoid saponins. Background Technology

[0002] Ginsenosides are the main active ingredients of ginseng-derived traditional Chinese medicines, possessing various activities such as anti-inflammatory, anti-tumor, and nerve cell protection. Ginsenosides Re and Rg1 are common active ingredients in ginseng, Panax notoginseng, and American ginseng, among other ginseng-derived traditional Chinese medicines. Rare ginsenosides Rh1 and F1 are intermediates in the synthesis of ginsenosides Re and Rg1, and are also important pharmacologically active components produced during the processing of ginseng and other traditional Chinese medicines.

[0003] Currently, the acquisition of ginsenosides mainly relies on plant extraction. This method is wasteful of precious Chinese medicinal materials, has high extraction costs, and only yields saponin components with relatively high content. The resource problem of rare components in precious Chinese medicinal materials is an important issue that needs to be addressed by traditional Chinese medicine. Summary of the Invention

[0004] In view of the shortcomings of the prior art described above, the present invention discloses a glycosyltransferase that can use protopanaxadiol as a substrate to achieve the biosynthesis of ginsenosides Rg1 and Re or rare ginsenosides Rh1, F1 and Rg2 through different enzyme combinations or whole-cell catalysis, thereby realizing the green manufacturing of rare ginsenosides.

[0005] The first aspect of the present invention is to provide a glycosyltransferase a, which is a protein having the following characteristics (a) or (b):

[0006] (a) A protein consisting of the amino acid sequence shown in SEQ ID NO.1;

[0007] (b) A protein derived from (a) whose amino acid sequence in (a) has been substituted, deleted or added with one or more amino acids and has glycosyltransferase activity.

[0008] Furthermore, the glycosyltransferase a is used to catalyze the transfer reaction of saponins from glucose to xylose. It can be used to catalyze reactions with different substrates.

[0009] Another aspect of the present invention provides a polynucleotide for encoding the aforementioned glycosyltransferase a.

[0010] Furthermore, the nucleotide sequence of the polynucleotide is shown in SEQ ID NO.3.

[0011] Furthermore, the amplification primers for the polynucleotide are shown in SEQ ID NO.7 and SEQ ID NO.8.

[0012] Another aspect of the present invention provides a recombinant plasmid containing the aforementioned polynucleotides.

[0013] Another aspect of the present invention provides a bioengineered bacterium containing the aforementioned recombinant plasmid. The bioengineered bacterium may be *Escherichia coli*.

[0014] Another aspect of the present invention provides that the above-mentioned glycosyltransferase a is used in any of the following reactions:

[0015] a. Catalyzing the reaction of triterpenoids with UDPG or UDPX to generate triterpenoids containing glucose or xylene at the C6 position.

[0016] Glycosylated triterpenoid saponins;

[0017] b. Catalyze the reaction of triterpenoids with UDPG or UDPX to generate triterpenoids with C20 hydroxyglucose or xylose.

[0018] For example, catalyzing the reaction of ginsenosides PPT and F1 with UDPG or UDPX to generate C6-glucose or xylosylated triterpenoid saponins, or catalyzing the reaction of ginsenoside Rg3 with UDPG or UDPX to generate corresponding saponins with C20-hydroxyglucose or xylosylation.

[0019] Furthermore, the triterpenoid compound may be selected from any of the following compounds:

[0020]

[0021] Another aspect of the present invention provides a glycosyltransferase composition containing glycosyltransferase a as described above.

[0022] Furthermore, the glycosyltransferase composition also contains glycosyltransferase b, which is a protein having the following characteristics (a) or (b):

[0023] (a) A protein consisting of the amino acid sequence shown in SEQ ID NO.2;

[0024] (b) A protein derived from (a) whose amino acid sequence in (a) has been substituted, deleted or added with one or more amino acids and has glycosyltransferase activity.

[0025] Another aspect of the present invention provides the application of the above-mentioned glycosyltransferase a, polynucleotide, recombinant plasmid, bioengineered bacteria, or glycosyltransferase composition in the preparation of triterpenoid saponins. This type of reaction refers to the reaction of reactants such as protopanaxadiol, ginsenoside F1, octetrazol, cycloastragalool, ginsenoside Rg3, etc., with UDPG or UDPX to generate the corresponding glycosylated triterpenoid saponins.

[0026] For example, glycosyltransferase a is first used to catalyze the reaction of PPT and UDPG to generate ginsenoside Rh1. Then, glycosyltransferase b is used to catalyze the reaction of ginsenoside Rh1 with uridine diphosphate rhamnose (UDP-rhamnose, UDPR) to generate ginsenoside Rg2.

[0027] For example, using α as a catalyst to catalyze the reaction of ginsenoside F1 with UDPG can generate ginsenoside Rg1.

[0028] For example, the application refers to using glycosyltransferase b to catalyze the reaction of ginsenoside Rg1 with UDPR to generate ginsenoside Re.

[0029] For example, the application refers to using glycosyltransferase a to catalyze the reaction of ginsenoside Rg3 with UDPG to generate ginsenoside Rd.

[0030] Furthermore, the catalytic temperature of the glycosyltransferase a is 35-45℃, preferably 40℃.

[0031] Furthermore, the catalytic temperature of the glycosyltransferase b is 35-45°C, preferably 40°C.

[0032] As described above, the application of UGT71A3 of the present invention as a catalyst in the preparation of triterpenoid saponins has the following beneficial effects:

[0033] Research has found that UGT71A3 has a highly efficient catalytic effect in the preparation of triterpenoid saponins, providing a new technical means for the preparation of rare triterpenoid saponins. Combining UGT71A3 with UGT94M1 can yield a wider variety of rare triterpenoid saponins. Attached Figure Description

[0034] Figure 1 Results of RNA electrophoresis examination of different Panax notoginseng samples.

[0035] Figure 2 Biosynthetic routes of ginsenosides Re and Rg1.

[0036] Figure 3 Biosynthetic route of ginsenoside Rd.

[0037] Figure 4 Chromatograms of the synthesis of ginsenosides Re and Rg2 catalyzed by PnUGT94M1.

[0038] Figure 5 Chromatogram of PnUGT71A3 catalyzing the synthesis of ginsenosides Rh1 and Rd.

[0039] Figure 6Thin-layer chromatography of PnUGT94M1 catalyzing the synthesis of Rg2 from Rh1 and the synthesis of Re from Rg1. Specifically, 1. Rg1, 2. PnUGT94M1 catalyzing the conversion of Rg1 to Re, 3. Re, 4. Rh1, 5. PnUGT94M1 catalyzing the conversion of Rh1 to Rg2, 6. Rg2;

[0040] Figure 7 Thin-layer chromatography of PnUGT71A3 catalyzing the synthesis of Rh1 from PPT and the synthesis of Rd from Rg3. Specifically, 1. Rg3, 2. UGT71A3 catalyzing the conversion of Rg3 to Rd, 3. Rd, 4. PPT, 5. UGT71A3 catalyzing the conversion of PPT to Rh1, and 6. Rh1;

[0041] Figure 8 Thin-layer chromatography of PnUGT71A3-catalyzed saponin glucosylation and xylosylation reactions. Specifically, 1-2. PnUGT71A3-catalyzed PPT and F1 glucosylation; 3-4. PnUGT71A3-catalyzed PPD and Rh2 glucosylation (no reaction); 5-6. PnUGT71A3-catalyzed Rg3 glucosylation and xylosylation; 7. Rd;

[0042] Figure 9 PnUGT71A3 catalyzes the C6-hydroxyl glycosylation of triterpenoid saponins. Specifically, 1. Rh1, 2. PnUGT71A3 catalyzes the PPT glycation reaction, 3. PPT, 4. Panaxtriol (PT), 5. PnUGT71A3 catalyzes the PT glycation reaction, 6. Cycloastragenol (CAG), 7. PnUGT71A3 catalyzes the CAG glycation reaction, 8. PnUGT71A3 catalyzes the F1 glycation reaction, and 9. Rg1.

[0043] Figure 10 Gel electrophoresis image after PCR amplification. In the image: 1. PnUGT71A3, 2. PnUGT71A1, 3. Marker, 4. PnUGT94M1. Detailed Implementation

[0044] Studies have found that glycosyltransferase a, namely UGT71A3, has the following amino acid sequence as shown in SEQ ID NO.1: MKNSELVFVPAPAIGHLVSMVEMAKLFISRHENLSITVLIAKFPIDTGINNYFESLLTNPTPRLTIVILPETEPQTYLNKPRHSILTALIESQKPHTRDIISGITRSESTRLVGLVADLLCVNIIDIANEFNVPTYVYSPAGAGFLGMVFHLQTLRDDENQDLTEFRNSDTELLVPSFENPVPAEVVPLMYLDKDGGSDLLLSMVRRYRETKGIIVNTFEELESYAINSLQSDSMIPQIYSVGPI LNLNGEGQNSDEAAVILGWLDDQPPSSVVFLCFGSFGSFQENQVKEIAIALERSGHNFLWSLRPRMPDGETKLQLKYSNLEEILPTGFLERTTSVGKVIGWAPQVAVLSHVAVGGFISHCGWNSTLESVWFGVPIATWPIYAEQQLNAFEMVKELGLAVEIEVDYKNEFNMKSDVIIRADEIETKIRKLMMDGKSNEIRTKVKEMKEKSRVAMAKNGSSYISLAKLIEEIM, has the ability to catalyze the reaction of glycosyl groups with aglycones to produce triterpenoid saponins. For example, it catalyzes the reaction of PPT with uridine diphosphate glucose (UDPG) to produce ginsenoside Rh1.

[0045] Research has shown that glycosyltransferase a is used in any of the following reactions:

[0046] a. Catalyzing the reaction of triterpenoids with UDPG or UDPX to generate triterpenoid saponins that glycosylate the C6 position of triterpenoids with glucose or xylose;

[0047] b. Catalyze the reaction of triterpenoids with UDPG or UDPX to generate triterpenoids with C20 hydroxyglucose or xylose.

[0048] For example, the glycosyltransferase a catalyzes the reaction of PPT and UDPG to generate ginsenoside Rh1, and the catalyst catalyzes the reaction of ginsenoside F1 and UDPG to generate ginsenoside Rg1.

[0049] The thin-layer chromatograms obtained from the specific reaction are as follows: Figures 7-8 .

[0050] For example, glycosyltransferase a can also be used to catalyze the following reactions:

[0051]

[0052] For example, PnUGT71A3 can be used to catalyze the glycation of PPT, the glycosylation of CAG, and the glycosylation of ginsenoside F1. The specific thin-layer chromatograms obtained from these reactions are shown below. Figure 9 .

[0053] In addition, in this article Figure 2 and Figure 3 It also shows some of the chemical reactions involved by glycosyltransferase a.

[0054] Furthermore, glycosyltransferase b, namely UGT94M1, was used, with the amino acid sequence shown in SEQ ID NO.2: MDTNEKTRIKVVMLPWLAYGHISPYLELAKKLSKRDFYVYFCSTSINLSSIRKKLAVDDHDAIQLIEFQLTSQTELPPHHHTTKGLPPHLIPDLIKALGLSGPNVINILNTVNPDLIIYDVFQLWVPAFAASLQIPAVHFQVVGALSTAAVYRFKVDSSIPVPCSRIFLDDTNIRKSPDYDSSSGENSGIVDLTFGTAIQSSDIILIKSSREFDEKNIEYYSLLMDKKKIVPTGPLVQVNTCVAVHTENEKD DIMDWLSKKEESSTVYVSFGSECYLSEPRIRELAHGLELSNVNFIWVISFPEGDEEMCNTSIEDVLPEGFLDRVKDRGVIVSWAPQERILGHGGLGGFVSHCGWGSVVEAMSYGVPIIAMPAQYEQPLHAMFVEEVGVGVEVLKDESGEFRRDEIAKAIKKVVVEKNGEGVRKKAREMGKAIKKRGEEEVECVVEELTKLCKKYQKVAAGQGKEWS, which can also catalyze the reaction of ginsenoside Rh1 with uridine diphosphate rhamnose (UDPR) to generate ginsenoside Rg2.

[0055] Using PPT as a raw material and PnUGT71A1 as a catalyst, ginsenoside F1 can be generated by reacting with UDPG. Glycosyltransferase a catalyzes the reaction of ginsenoside F1 with UDPG to generate ginsenoside Rg1. Glycosyltransferase b catalyzes the reaction of ginsenoside Rg1 with UDPR to generate ginsenoside Re.

[0056] Glycosyltransferase a catalyzed reaction of ginsenoside Rg3 with UDPG to generate ginsenoside Rd.

[0057] Both glycosyltransferase a and glycosyltransferase b act as catalysts for the above reactions, resulting in high yields.

[0058] Following further research and exploration, this application provides a polynucleotide encoding glycosyltransferase a, the nucleotide sequence of which is shown in SEQ ID NO.3:

[0059]

[0060] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention. It should be noted that the process equipment or apparatus not specifically specified in the following embodiments are all conventional equipment or apparatus in the art. Furthermore, it should be understood that one or more method steps mentioned in the present invention do not exclude the existence of other method steps before or after the combined steps, or the insertion of other method steps between these explicitly mentioned steps, unless otherwise stated; it should also be understood that the combined connection relationship between one or more devices / apparatus mentioned in the present invention does not exclude the existence of other devices / apparatus before or after the combined devices / apparatus, or the insertion of other devices / apparatus between these explicitly mentioned two devices / apparatus. Moreover, unless otherwise stated, the numbering of each method step is only a convenient tool for identifying each method step, and not for limiting the order of the method steps or limiting the scope of the present invention. Changes or adjustments to their relative relationships, without substantially altering the technical content, should also be considered within the scope of the present invention.

[0061] Ginsenoside Re: CAS No.: 52286-59-6 Molecular Formula: C 48 H 82 O 18 Molecular weight: 947.15;

[0062] Ginsenoside Rg1: CAS No.: 22427-39-0 Molecular Formula: C 42 H 72 O 14 Molecular weight: 801.01;

[0063] Ginsenoside Rh1: CAS No.: 63223-86-9 Molecular Formula: C 36 H 62 O9 molecular weight: 638.87;

[0064] Ginsenoside F1: CAS No.: 53963-43-2; Molecular formula: C36H62O9; Molecular weight: 638.88;

[0065] Ginsenoside Rg2: CAS No.: 52286-74-5 Molecular Formula: C 42 H 72 O 13 Molecular weight: 785.01;

[0066] Ginsenoside Rd: CAS No.: 52705-93-8 Molecular Formula: C 48 H 82 O 18 Molecular weight: 947.17.

[0067] Material:

[0068] LB liquid medium: peptone 10 g / L, yeast extract 5 g / L, sodium chloride 10 g / L.

[0069] LB solid medium: Add 15 g / L agarose to LB liquid medium.

[0070] HPLC-MS detection method: Mass spectrometry conditions in both positive and negative ion modes were optimized according to the molecular weight of the compounds to be detected. The compounds to be detected are shown in Table 1. Mass spectrometry parameters: Interface temperature: 300℃, DL temperature: 300℃, Heat block temperature: 400℃, Drying gas flow: 10L / min, Heating gas flow: 10L / min, Nebulizing gas flow: 2.5L / min.

[0071] Table 1 List of Detected Compounds

[0072]

[0073]

[0074] Chromatographic column: Waters Acquity BEH C18, 1.7 μm, 2.1 × 50 mm;

[0075] Mobile phase A: 0.1% formic acid aqueous solution; Mobile phase B: acetonitrile;

[0076] Flow rate: 0.3 ml / min; Column temperature: 45℃; Elution: 0-1 min 30% B; 1-2 min 30-40% B;

[0077] 2-4min 40-80%B; 4-6min 80%B; 6-6.01min 80-30%B; 6.01-8min 30%B.

[0078] Example 1: Cloning of a glycosyltransferase gene derived from Panax notoginseng

[0079] Fresh Panax notoginseng plants were harvested and rinsed twice with deionized water to remove attached soil and microorganisms. Different tissues (roots, stems, and leaves) were separated and flash-frozen in liquid nitrogen. Total RNA was extracted from the different tissues using the TRIzol method, and cDNA was synthesized using a rapid cDNA reverse transcription kit. The cDNA was amplified using primers listed in Table 1 as a template. The amplified products were subjected to gel electrophoresis, and the results are shown below. Figure 10 The target sequence was confirmed. The obtained fragment was digested with enzymes and ligated into the pMAL-c2x vector, transformed into E. coli DH5α, and positive clones were selected for sequencing.

[0080] Table 2

[0081]

[0082] Example 2 Construction of Recombinant Glycosyltransferase Expression Vector and Recombinant Engineered Escherichia coli

[0083] Select the positive clones with correct sequencing, culture the bacteria, extract the plasmid, and transfer the plasmid into the protein expression strain to construct the Rosetta(DE3) engineered bacteria.

[0084] Example 3 Expression of recombinase

[0085] The engineered bacteria were cultured at 37℃ for 3-4 hours (OD). 600 To achieve a concentration of approximately 0.8, add 0.35 mL of 1M IPTG to 1 L of bacterial culture and continue induction at 16℃ for 18-24 h. After induction, collect the bacterial cells by centrifugation at 4000g, wash once with 100 mL of 50 mM pH 7.5 Tris-HCl, centrifuge again, add 100 mL of lysis buffer (100 mM pH 7.5 Tris-HCl, 200 mM NaCl, 10% glycerol, 1 mM PMSF), homogenize by high pressure, and centrifuge to obtain the supernatant containing the glycosyltransferase (PnUGT71A3).

[0086] The supernatant of glycosyltransferase PnUGT71A1 was prepared using the same method.

[0087] The supernatant of glycosyltransferase PnUGT94M1 was prepared using the same method.

[0088] The supernatant of sucrose synthase (AtSUS) was prepared using the same method. Sucrose synthase can catalyze the reaction of uridine diphosphate (UDP) with sucrose to produce UDPG. By combining glycosyltransferase with sucrose synthase and adding sucrose and a catalytic amount of UDP, in-situ synthesis and regeneration of UDPG can be achieved, thus avoiding the direct use of expensive UDPG.

[0089] Example 4: UGT71A3 catalyzes the production of ginsenoside Rh1 from PPT.

[0090] Add 50 mL each of the supernatant from glycosyltransferase PnUGT71A3 and sucrose synthase AtSUS1 to 50 mL of 500 mM pH 7.5 phosphate buffer, then add ginsenoside PPT to a final concentration of 1 g / L, sucrose to a final concentration of 20 g / L, and UDP to a final concentration of 0.2 g / L. Finally, add water to a total reaction volume of 300 mL (the obtained HPLC chromatogram is shown below). Figure 5 At a temperature of 40℃ and a rotation speed of 220 rpm, samples were taken for analysis at 2h, 8h, 14h, and 20h. When the PPT conversion rate reached over 90%, an equal volume of n-butanol was added to terminate the reaction, and saponins were extracted. This process was repeated twice. The extracts were combined, dried under reduced pressure, and purified using silica gel. Thin-layer chromatography was then performed to obtain the desired product. Figure 7 The chromatogram yielded approximately 0.348 g of ginsenoside Rh1 with a purity greater than 98% and a molar yield greater than 85%. The chemical reaction equation is shown below. Figure 2 .

[0091] Example 5: UGT71A3 combined with UGT71A1 catalyzes the production of ginsenoside Rg1 from PPT.

[0092] 50 mL each of the supernatants of glycosyltransferases PnUGT71A3, PnUGT71A1, and sucrose synthase AtSUS1 were added to 50 mL of 500 mM pH 7.5 phosphate buffer. Ginsenoside PPT was added to a final concentration of 1.0 g / L, sucrose to a final concentration of 30 g / L, and UDP to a final concentration of 0.25 g / L. Water was added to a total reaction volume of 300 mL. The reaction was carried out at 40℃ and 220 rpm for 2 h, 8 h, 14 h, and 20 h. When the conversion of PPT to Rg1 reached over 80% and there was virtually no residue of intermediates Rh1 and F1, an equal volume of n-butanol was added to terminate the reaction, and the saponins were extracted. This process was repeated three times. The extracts were combined, dried under reduced pressure, and purified using silica gel to obtain approximately 0.409 g of ginsenoside Rg1 with a purity greater than 98% and a molar yield of 81%. The chemical reaction equation is shown below. Figure 2 .

[0093] Example 6: Biosynthesis of ginsenoside Rd catalyzed by PnUGT71A3

[0094] Add 50 mL each of the supernatant from glycosyltransferase PnUGT71A3 and sucrose synthase AtSUS1 to 50 mL of 500 mM pH 7.5 phosphate buffer, then add ginsenoside Rg3 to a final concentration of 1 g / L, sucrose to a final concentration of 20 g / L, and UDP to a final concentration of 0.2 g / L. Add water to a total reaction volume of 300 mL. (The obtained HPLC chromatogram is shown below.) Figure 5 At a temperature of 40℃ and a rotation speed of 220 rpm, samples were taken for analysis at 2h, 8h, 14h, and 20h. The reaction was continued until the Rg3 spot essentially disappeared. An equal volume of n-butanol was added to terminate the reaction, and the saponins were extracted. This process was repeated twice. The extracts were combined, dried under reduced pressure, and purified using silica gel. Thin-layer chromatography was then performed to obtain the desired product. Figure 7 The chromatogram yielded approximately 0.301 g of ginsenoside Rd with a purity greater than 98% and a molar yield of 83%. The chemical reaction equation is shown below. Figure 3 .

[0095] Example 7: Biosynthesis of Ginsenoside Rg2 catalyzed by PnUGT94M1

[0096] Add 50 mL of the supernatant of glycosyltransferase PnUGT94M1 to 50 mL of 500 mM pH 7.5 phosphate buffer, add ginsenoside Rh1 to a final concentration of 1 g / L, add UDP-rhamnose to a final concentration of 1.2 g / L, and add water to a total reaction volume of 200 mL (the obtained HPLC chromatogram is shown below). Figure 4 At a temperature of 40℃ and a rotation speed of 220 rpm, samples were taken for analysis at 2h, 8h, 14h, and 20h. When the Rh1 conversion reached over 90%, an equal volume of n-butanol was added to terminate the reaction, and the saponins were extracted. This process was repeated twice, and the extracts were combined, dried under reduced pressure, and purified using silica gel. Thin-layer chromatography was then performed to obtain the desired product. Figure 6 The chromatogram yielded approximately 0.185 g of ginsenoside Rg2 with a purity greater than 98% and a molar yield greater than 75%. For specific reaction details, please refer to [reference needed]. Figure 2 .

[0097] Example 8: Biosynthesis of Ginsenoside Re catalyzed by recombinase

[0098] Add 50 mL of the supernatant of glycosyltransferase PnUGT94M1 to 50 mL of 500 mM pH 7.5 phosphate buffer, add ginsenoside Rg1 to a final concentration of 1 g / L, add UDP-rhamnose to a final concentration of 1.2 g / L, and add water to a total volume of 200 mL. Incubate at 40℃ (the obtained HPLC chromatogram is shown below). Figure 4At a rotation speed of 220 rpm, samples were taken for analysis at 2 h, 8 h, 14 h, and 20 h. When the conversion of Rg1 reached over 90%, an equal volume of n-butanol was added to terminate the reaction, and saponins were extracted. This process was repeated twice. The extracts were combined, dried under reduced pressure, and purified using silica gel. Thin-layer chromatography was then performed to obtain the desired product. Figure 6 The chromatogram yielded approximately 0.189 g of ginsenoside Re with a purity greater than 98% and a molar yield greater than 80%. For specific reaction details, please refer to [reference needed]. Figure 2 .

[0099] The above embodiments are for illustrating the implementation schemes disclosed in this invention and should not be construed as limiting the invention. Furthermore, various modifications listed herein, as well as variations in the methods and compositions of the invention, will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been specifically described in conjunction with various specific preferred embodiments, it should be understood that the invention should not be limited to these specific embodiments. In fact, various modifications as described above that are obvious to those skilled in the art to obtain the invention should be included within the scope of this invention.

Claims

1. A glycosyltransferase a, characterized in that, The amino acid sequence of the glycosyltransferase a is shown in SEQ ID NO.

1.

2. A polynucleotide, characterized in that: The polynucleotide is used to encode the glycosyltransferase a described in claim 1.

3. The polynucleotide according to claim 2, characterized in that: The nucleotide sequence of the polynucleotide is shown in SEQ ID NO.

3.

4. A recombinant plasmid, characterized in that: The plasmid contains the polynucleotide as described in claim 2.

5. A type of bioengineered bacterium, characterized in that: The bioengineered bacteria contain the recombinant plasmid as described in claim 4.

6. A glycosyltransferase composition, characterized in that, The composition contains the glycosyltransferase a as described in claim 1.

7. The glycosyltransferase composition according to claim 6, characterized in that: The glycosyltransferase composition further contains glycosyltransferase b, which is a protein composed of the amino acid sequence shown in SEQ ID NO.

2.

8. The application of the glycosyltransferase a as described in claim 1, or the polynucleotide as described in any one of claims 2-3, or the recombinant plasmid as described in claim 4, or the bioengineered bacteria as described in claim 5 in catalyzing the reaction of protopanaxadiol with UDPG to generate ginsenoside Rh1, or the reaction of ginsenoside F1 with UDPG to generate ginsenoside Rg1, or the reaction of ginsenoside Rg3 with UDPG to generate ginsenoside Rd.

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