A glycosyltransferase mutant and its application in the synthesis of salidroside
By directed evolution of the glycosyltransferase UGTBS from Bacillus subtilis 168, the efficient mutant UGTBS-M4 was obtained, which solved the problem of low expression and contamination in rhodioside extraction, and achieved efficient and green biosynthetic rhodioside, which was suitable for industrial production.
Patent Information
- Application Number
- CN202510062199.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-15
- Publication Date
- 2025-07-25
- Estimated Expiration
- 2045-01-15
AI Technical Summary
The extraction method of rhodiolide in the prior art has the problem of low expression of plant-derived glycosyltransferases and polluting the environment by chemical synthesis, making it difficult to achieve efficient and green industrial production.
By directed evolution of the glycosyltransferase UGTBS derived from Bacillus subtilis 168, the mutant UGTBS-M4 was obtained, which improved its soluble expression and catalytic efficiency in E. coli, and the biosynthesis method was used to catalyze the synthesis of rhodiosides of tyrosol.
The high conversion rate and high yield of rhodiolisin were achieved. The substrate conversion rate of tyrosin synthesis of rhodiolisin catalyzed by the mutant UGTBS-M4 was 95%, the yield reached 55.0 g/L, and the spatiotemporal yield was 1.3 g/L/h, which solved the problems of low expression and contamination in the prior art.
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Abstract
Description
Technical Field
[0001] The present invention belongs to the field of enzyme engineering and relates to a glycosyltransferase mutant and its application in the synthesis of salidroside. Background Art
[0002] Salidroside, also known as rhodioloside, has the molecular formula C 14 H 20 O7 and a molecular weight of 300.30. It is an important active ingredient in the traditional Chinese medicine Rhodiola rosea and has various pharmacological activities such as antioxidant, anti-inflammatory, anti-tumor, anti-cardiovascular disease, and anti-aging effects, showing broad application prospects. Salidroside is a glycoside compound synthesized with tyrosol (p-hydroxyphenethyl alcohol) as the aglycone and glucose as the glycosyl donor.
[0003] Currently, the extraction methods of salidroside mainly include plant extraction method and chemical synthesis method. Extracting from Rhodiola plants faces problems such as low content, high extraction cost, and complex processes. The chemical synthesis process requires selective protection of the substrate hydroxyl group, and various reagents used in it will cause environmental pollution, which does not conform to the concept of environmental protection and is not conducive to industrial production.
[0004] Preparing glycoside compounds by biosynthesis is one of the key research directions in recent years. The enzyme-catalyzed biosynthesis method has attracted much attention due to its advantages such as simple operation, mild conditions, few by-products, high yield, and high efficiency, green and environmental protection. Salidroside is mainly obtained by glycosylation modification of tyrosol by glycosyltransferase. Through continuous exploration and discovery by relevant scientists, glycosyltransferases (UGT73B6, UGT72B14, UGT85A1) for synthesizing salidroside with tyrosol as the substrate have been discovered and identified, and the heterologous expression of these glycosyltransferases and the synthesis of salidroside have been achieved through microorganisms such as Escherichia coli. However, the low expression level of plant-derived glycosyltransferases limits the synthesis of salidroside. Summary of the Invention
[0005] To solve the above problems, the present invention provides a glycosyltransferase mutant and its application in the synthesis of salidroside. By performing directed evolution based on the protein structure on glycosyltransferase UGT BS efficient mutants are successfully obtained, the yield of the biocatalytic preparation of salidroside from tyrosol is improved, and a method for biosynthesizing salidroside is provided. This method has a high conversion rate of salidroside and is suitable for the industrial production of salidroside.
[0006] Our laboratory has discovered glycosyltransferase UGT BS from Bacillus subtilis 168. This enzyme can be highly expressed in a soluble form in Escherichia coli and has the activity of catalyzing the synthesis of salidroside from tyrosol in the presence of uridine diphosphate glucose (UDPG).
[0007] The enzyme molecule is modified using the theory of directed evolution to obtain a glycosyltransferase mutant with high catalytic efficiency. Based on the protein structure of glycosyltransferase and ligands, a simulation docking experiment is carried out in the present invention to obtain a complex model of the enzyme and the ligand. Based on the rational design of the protein structure, the active site is found by analyzing the protein crystal structure and mutated to obtain a glycosyltransferase mutant for highly efficient synthesis of salidroside, which is of great significance for the industrial production of salidroside.
[0008] The present invention uses the glycosyltransferase UGT derived from Bacillus subtilis 168 previously discovered in the laboratory BS as a template (the amino acid sequence is shown in SEQ ID NO.1), and site-directed mutagenesis is carried out on it to obtain the mutant UGT BS -M4.
[0009] The present invention provides a glycosyltransferase mutant, which is obtained by simultaneously mutating the 62nd, 129th, 168th and 316th amino acids on the basis of the amino acid sequence of the glycosyltransferase UGT BS shown in SEQ ID NO:1.
[0010] In one embodiment of the present invention, the mutant is based on the glycosyltransferase UGT BS shown in SEQ ID NO:1, where the isoleucine at the 62nd position is mutated to asparagine, the serine at the 129th position is mutated to threonine, the phenylalanine at the 168th position is mutated to tryptophan, and at the same time the tyrosine at the 316th position is mutated to serine, and it is named UGT BS -M4.
[0011] In one embodiment of the present invention, the amino acid sequence of the glycosyltransferase mutant UGT BS -M4 is shown in SEQID NO:2.
[0012] The present invention also provides a gene encoding the above-mentioned glycosyltransferase mutant UGT BS -M4.
[0013] The present invention also provides a recombinant vector carrying the above gene.
[0014] In one embodiment of the present invention, the recombinant vector uses pET-28a(+) as the expression vector.
[0015] The present invention also provides a recombinant cell expressing the above-mentioned glycosyltransferase mutant, or containing the above gene, or containing the above recombinant vector.
[0016] In one embodiment of the present invention, the recombinant cell uses bacteria or fungi as the expression host.
[0017] The present invention also provides a method for catalytically synthesizing salidroside. The method is to add the glycosyltransferase mutant UGT BS -M4, or the recombinant cell, to a reaction system containing tyrosol, and react to prepare salidroside.
[0018] In one embodiment of the present invention, the reaction system contains 1-10 mmol / L tyrosol, 2-20 mmol / L UDPG, 1-10% (v / v) DMSO, 5-40% (v / v) crude enzyme solution of glycosyltransferase mutant (the enzyme concentration in the reaction system is 0.1-1 mg / ml), and 50-200 mmol / L Tris-HCl buffer solution.
[0019] In one embodiment of the present invention, the reaction system contains 1-30 mmol / L tyrosol, 0.2-20 mmol / L UDP, 50-200 mmol / L Tris-HCl buffer solution, 400-1000 mmol / L sucrose, 30-40% (v / v) crude enzyme solution of glycosyltransferase mutant, and 15-30% (v / v) crude enzyme solution of sucrose synthase.
[0020] In one embodiment of the present invention, the reaction conditions are as follows: reacting at 30-40 °C for 0-50 h.
[0021] The present invention also discloses a catalyst containing the aforementioned glycosyltransferase mutant, or the aforementioned gene, or the aforementioned recombinant cell.
[0022] Beneficial effects:
[0023] Compared with the salidroside glycosyltransferase from plants, the microbial salidroside glycosyltransferase discovered in the present invention is more easily expressed efficiently and solubly in model strains such as Escherichia coli;
[0024] The glycosyltransferase mutant obtained in the present invention has a significantly improved glycosylation activity towards tyrosol. The highest titer of salidroside synthesized by using the glycosyltransferase mutant to catalyze tyrosol can reach 55.0 g / L, the substrate conversion rate is 97.9%, and the space-time yield is 1.3 g / L / h.
[0025] Experimental results confirm that the present invention can promote the industrial production of salidroside by biosynthesis, successfully solve the key problem of low glycosyltransferase activity in the enzymatic preparation process of salidroside, and the obtained mutant shows great industrial application prospects. Description of the Drawings
[0026] Figure 1For the glycosyltransferase mutant UGT BS -M4 protein electrophoresis pattern, where M is Marker, 1 is the unpurified protein, and 2 is the purified protein;
[0027] Figure 2 In the single-enzyme system, wild-type UGT BS and the mutant UGT BS -M4 HPLC chromatogram of catalyzing tyrosol to produce salidroside;
[0028] Figure 3 In the double-enzyme system, wild-type UGT BS and the mutant UGT BS -M4 schematic diagram of catalytic efficiency and regioselectivity. Detailed implementation manners
[0029] The present invention will be further described below in conjunction with the specification drawings and specific embodiments, but the embodiments do not impose any form of limitation on the present invention. Unless otherwise specified, the methods and equipment used in the present invention are conventional methods and equipment in the technical field.
[0030] Unless otherwise specified, the kit materials used in the following embodiments are all commercially available products.
[0031] The experimental methods described in the present invention are all conventional methods unless otherwise specified.
[0032] The DNA polymerase and DpnⅠ enzyme used in the embodiments of the present invention are purchased from Takara Biotechnology Co., Ltd., and the recombinant cloning kit and plasmid extraction kit are both purchased from Axygen Biotechnology Co., Ltd. The primer synthesis and gene sequencing work are completed by General Biology (Anhui) Co., Ltd.
[0033] The tyrosol and UDPG used in the present invention are both purchased from Shanghai Macklin Biochemical Co., Ltd.
[0034] The expression vector used in the present invention is pET-28a(+), and the host used is Escherichia coli BL21(DE3).
[0035] Example 1 Cloning of the glycosyltransferase UGT BS gene
[0036] Using the Bacillus subtilis 168 genome as a template, F and R primers were designed as shown in Table 1. The F primer is SEQ ID NO:3, and the R primer is SEQ ID NO:4. The wild-type glycosyltransferase gene was amplified, and its encoded amino acid sequence is as shown in SEQ ID NO.1, named UGT BS, the amplified gene was ligated to the pET-28a(+) vector using a one-step cloning kit (ClonExpressⅡ One Step Cloning Kit) to obtain the recombinant plasmid pET28a-UGT BS . The recombinant plasmid was transformed into E.coil BL21(DE3), and the transformation products were spread on LB solid medium containing kanamycin and cultured at 37 °C for 12 - 16 h. Transformants were picked, verified by colony PCR, and positive clones were selected and sent for sequencing to obtain the recombinant engineering bacteria containing the UGT BS gene.
[0037] Table 1 Primers used for cloning UGT BS
[0038]
[0039] Note: Lowercase bases are the parts that recombine and ligate with the vector
[0040] SEQ ID NO.1 UGT BS
[0041] MKKYHISMINIPAYGHVNPTLALVEKLCEKGHRVTYATTEEFAPAVQQAGGE
[0042] ALIYHTSLNIDPKQIREMMEKNDAPLSLLKESLSILPQLEELYKDDQPDLIIYDF
[0043] VALAGKLFAEKLNVPVIKLCSSYAQNESFQLGNEDMLKKIREAEAEFKAYLEQ
[0044] EKLPAVSFEQLAVPEALNIVFMPKSFQIQHETFDDRFCFVGPSLGERKEKESLLI
[0045] DKDDRPLMLISLGTAFNAWPEFYKMCIKAFRDSSWQVIMSVGKTIDPESLEDIP
[0046] ANFTIRQSVPQLEVLEKADLFISHGGMNSTMEAMNAGVPLVVIPQMYEQELT
[0047] ANRVDELGLGVYLPKEEVTVSSLQEAVQAVSSDQELLSRVKNMQKDVKEAG
[0048] GAERAAAEIEAFMKKSAVPQ
[0049] Induced Expression of Recombinant Strain in Example 2 and Purification of Target Protein
[0050] The recombinant strain pET28a-UGT constructed in Example 1 BS was inoculated into 10 mL of LB liquid medium containing 50 μg / mL kanamycin and cultured overnight at 37 °C with 180 rpm. Then, it was transferred to 50 mL of fresh LB liquid medium containing kanamycin resistance at an inoculation amount of 2% and cultured at 37 °C with 200 rpm. When the OD 600 reached 0.6 - 0.8, isopropyl-β-D-thiogalactoside (IPTG) with a final concentration of 1 mM was added for induction at 28 °C with 200 rpm for 24 h. After the induction ended, cell precipitation was obtained by centrifugation at 4 °C, 5000 rpm for 20 min. After pouring out the supernatant, 25 mL of Tris-HCl (pH 8.0, 50 mM) buffer was added to suspend evenly. The cells were lysed with an ultrasonic cell disruptor and then centrifuged to collect the supernatant, obtaining the crude enzyme solution. SDS-PAGE analysis was performed to detect the protein expression, and the results were as Figure 1 shown. It can be seen that UGT BS can be highly efficiently and solubly expressed in Escherichia coli.
[0051] UGT was purified by nickel column affinity chromatography: The crude enzyme solution was obtained according to the method in Example 2, passed through a 0.45 μm filter membrane, and the sample was slowly loaded onto the column to allow the protein to fully bind to the nickel column; the impurity proteins were washed with 10 times the volume of lysis buffer, and the target protein was eluted with 10 times the volume of elution buffer. The eluate was collected and then dialyzed and stored at -80 °C. BS
[0052] Example 3 Determination of Glycosyltransferase Enzyme Activity: The enzyme activity was detected by analyzing the results of the glycosylation reaction by liquid phase
[0053] The glycosylation reaction was carried out in a 200 μL reaction system as follows: 50 mM Tris-HCl (pH 8.0), 2 mM tyrosol, 2% DMSO, 4 mM UDPG, and 40 μL of crude enzyme solution. The reaction was carried out at 35 °C with 1000 rpm for 1 h. The reaction was terminated by adding 4 times the volume of methanol, centrifuged at 12000 rpm for 1 min, passed through a 0.22 μm filter membrane, and then analyzed by HPLC. HPLC used an EclipsePlus C18 chromatographic column (4.6 mm × 150 mm, 5 μm diameter), the injection volume was 10 μL, the column temperature was 30 °C, the detection wavelength was 275 nm, and the mobile phase used A pipeline: water, B pipeline: methanol, and the flow rate was 0.8 mL / min. The specific program is shown in Table 2:
[0054] Table 2 HPLC Elution Program
[0055]
[0056] Enzyme activity definition: The amount of enzyme required to glycosylate 1 μmoL tyrosol per hour under standard assay conditions.
[0057] It was determined that the specific enzyme activity of the wild-type glycosyltransferase UGT BS was 3.6 U / mg.
[0058] Example 4 Site-directed saturation mutagenesis of wild-type glycosyltransferase UGT BS
[0059] In the present invention, amino acids near the substrate-binding pocket were screened out as mutation sites through homology modeling. These amino acids include G15, I62, V108, S129, L138, F168, F231, M315, Y316, E317, Q318, L320.
[0060] In the present invention, specific primers were designed (Table 3, SEQ ID NOs: 5-28 from top to bottom in sequence). Using the recombinant plasmid pET28a-UGT BS as a template and a pair of primers with mutation sites, PCR amplification was performed using Prime Star to obtain recombinant plasmids with specified mutation sites. The amplification products were digested with DpnⅠ digestion enzyme at 37 °C for 3 h to degrade the initial template. The digested products were transformed into E.coil BL21(DE3) and spread on a solid plate containing kanamycin and cultured overnight at 37 °C.
[0061]
[0062]
[0063]
[0064]
[0065] Table 3 Primers used for UGT BS mutants
[0066] Where N represents A, C, G or T; M represents A or C; K represents G or T.
[0067] Example 5 Screening of mutant libraries
[0068] On the mutant plates at each site, clone colonies were picked and transferred to a 96-well deep-well plate containing kanamycin and 1 mL of LB liquid medium for culture. When OD 600 When it reached 0.6 - 0.8, the cells were induced with 1 mM IPTG. After the induction, cell pellets were obtained by centrifugation at 4°C, 5000 rpm for 20 min. After pouring off the supernatant, the cell pellets were resuspended evenly with Tris-HCl (pH 8.0, 50 mM) buffer, 2 mM tyrosol, 4 mM UDPG, and 2% DMSO were added, and the reaction was carried out for 12 h. 4 volumes of methanol were added to terminate the reaction. The screening was carried out by the liquid phase method described in Example 3. Mutant strains with obvious effects were selected for plate transfer, induced expression using a 24-well plate, centrifuged, the precipitate was resuspended with 50 mM Tris-HCl, broken, 40 μL of crude enzyme solution was taken and reacted with 2 mM tyrosol, 4 mM UDPG, 2% DMSO, and 50 mM Tris-HCl (pH 8.0) for 12 h, and the catalytic effect was further verified by liquid phase. The selected good mutant strains were sequenced.
[0069] Example 6 Combinatorial Mutation
[0070] The single mutation sites with significantly improved activity in Example 5 were combined to obtain double mutants with different combination methods. Further, the mutants with significantly improved activity in the double mutation sites were combined with other high-activity sites for combinatorial mutation. Then, the crude enzyme solutions of each mutant combination were used in a 200 μL reaction system of 40 μL of crude enzyme solution, 2 mM tyrosol, 4 mM UDPG, 2% DMSO, and 50 mM Tris-HCl (pH 8.0), reacted at 35°C and 1000 rpm for 12 h, and 4 volumes of methanol were added to terminate the reaction. Detection was carried out by the liquid phase method described in Example 3.
[0071] Wild-type UGT BS to mutant UGT BS - Changes in substrate conversion rate and regioselectivity during the mutation process of M4:
[0072] As shown in Table 4, compared with the wild-type glycosyltransferase UGT BS the catalytic efficiency and regioselectivity of the glycosyltransferase mutant UGT BS -M4 have been greatly improved.
[0073] Among them, mutant M1 compared with the wild-type glycosyltransferase UGT BS , only the 62nd amino acid is mutated to N. Mutant M2 compared with the wild-type glycosyltransferase UGT BS , the 62nd amino acid is mutated to N, and the 129th amino acid is mutated to T. Mutant M3 compared with the wild-type glycosyltransferase UGT BS , the 62nd amino acid is mutated to N, the 129th amino acid is mutated to T, and the 168th amino acid is mutated to W.
[0074] Table 4 Substrate conversion rate and regioselectivity from single-point to four-point mutations
[0075]
[0076] The substrate conversion rate refers to: the total substrate concentration entering the reaction system minus the remaining substrate concentration after the reaction is completed, and then divided by the total substrate concentration entering the reaction system.
[0077] The regioselectivity refers to: the molar concentration ratio of salidroside in two products (salidroside and icariside D2).
[0078] After several rounds of iterative saturation mutagenesis, a glycosyltransferase mutant UGT BS -M4 with significantly improved activity was obtained, and its amino acid sequence is shown in SEQ ID NO:2. The wild-type glycosyltransferase UGT BS and the glycosyltransferase mutant UGT BS -M4 catalyzed tyrosol to obtain the product as shown in the liquid chromatography Figure 2 . The results show that compared with the wild-type glycosyltransferase UGT BS , the catalytic activity of the mutant UGT BS -M4 has been significantly improved, the substrate conversion rate has increased significantly, and the regioselectivity has also increased to a certain extent (that is, the by-product icariside D2 generated is significantly less, and the substrate selectivity of the mutant UGT BS -M4 for tyrosol is better than that of the wild-type glycosyltransferase UGT BS ).
[0079] In the single-enzyme catalytic system, the glycosylation activity of the glycosyltransferase mutant obtained in the present invention for tyrosol has been significantly improved, and the substrate conversion rate has increased from 44% of the wild-type glycosyltransferase UGT BS to 95% of the mutant UGT BS -M4. In addition, the proportion of salidroside in the two obtained products has increased from 82% of the wild-type to 98% of the mutant UGT BS -M4, and % is the molar concentration ratio.
[0080] In addition, the specific enzyme activity of the UGT BS -M4 mutant was measured by the method and system in Example 3. The specific enzyme activity of the mutant UGT BS -M4 is 7.3 U / mg. Compared with the wild-type UGT BS , the catalytic activity of the glycosyltransferase mutant UGT BS -M4 has been significantly improved.
[0081] SEQ ID NO.2 UGT BS -M4
[0082] MKKYHISMINIPAYGHVNPTLALVEKLCEKGHRVTYATTEEFAPAVQQAGGEA
[0083] LIYHTSLNNDPKQIREMMEKNDAPLSLLKESLSILPQLEELYKDDQPDLIIYDFV
[0084] ALAGKLFAEKLNVPVIKLCSTYAQNESFQLGNEDMLKKIREAEAEFKAYLEQE
[0085] KLPAVSWEQLAVPEALNIVFMPKSFQIQHETFDDRFCFVGPSLGERKEKESLLI
[0086] DKDDRPLMLISLGTAFNAWPEFYKMCIKAFRDSSWQVIMSVGKTIDPESLEDIP
[0087] ANFTIRQSVPQLEVLEKADLFISHGGMNSTMEAMNAGVPLVVIPQMSEQELTA
[0088] NRVDELGLGVYLPKEEVTVSSLQEAVQAVSSDQELLSRVKNMQKDVKEAGG
[0089] AERAAAEIEAFMKKSAVPQ
[0090] Example 7 Mutant Glycosyltransferase UGT BS - Preparation of Salidroside Using M4
[0091] For the best mutant UGT obtained in Example 6 BSThe -M4 strain was used for protein expression according to the method of Example 2 to obtain a crude enzyme solution. The sucrose synthase gene was AtSuSy from Arabidopsis thaliana (GenBank ID: NP_001031915.1). It was constructed into the pET-32a(+) vector to obtain a plasmid, and protein expression was carried out according to Example 2 to obtain a crude enzyme solution. The 10 mL catalytic reaction system was: 50 mM Tris-HCl (pH 8.0), 0.5 mM UDP (uridine diphosphate), 800 mM sucrose, 2% DMSO, 30 mM tyrosol, 3.42 mL crude glycosyltransferase enzyme solution and 2.18 mL crude sucrose synthase enzyme solution, and the reaction was carried out at 35 °C. 30 mM tyrosol was added at 0 h and 2 h respectively, and 20 mM tyrosol was added at 6 h, 9 h, 12 h, 15 h, 18 h, 21 h, and 24 h. At 7 h, 15 h, 21 h, 30 h, and 35 h, 3.42 mL crude glycosyltransferase enzyme solution and 2.18 mL crude sucrose synthase enzyme solution were respectively supplemented. In the presence of UDP, sucrose was catalyzed by sucrose synthase to produce fructose and UDPG, and the generated UDPG participated in the synthesis of salidroside again.
[0092] The results of the fed-batch reaction at 42 h are as Figure 3 shown. The substrate conversion rate of the wild-type glycosyltransferase UGT BS was 50.3%, the final yield of salidroside preparation was 22.0 g / L, the space-time yield was 0.5 g / L / h, the final yield of icariside D2 was 8.2 g / L, and the space-time yield was 0.2 g / L / h; the mutant glycosyltransferase UGT BS -M4 had a substrate conversion rate of 97.9%, the final yield of salidroside preparation was 55.0 g / L, the space-time yield was 1.3 g / L / h, the final yield of icariside D2 was 1.6 g / L, and the space-time yield was 0.04 g / L / h.
[0093] Based on the amino acid sequence of the glycosyltransferase shown in SEQ ID NO:1, the present invention carried out iterative saturation mutagenesis through molecular modification technology. The enzyme activity and catalytic efficiency of the modified glycosyltransferase mutant were greatly improved. Mutations at specific sites could significantly promote the glycosylation activity of the tyrosol substrate. The mutant enzyme was obtained by inducing expression and protein purification of the obtained mutant strain. The substrate conversion rate of the modified glycosyltransferase mutant catalyzing the tyrosol glycosylation reaction could reach more than 95%, and the yield reached 55.0 g / L, providing an efficient and green new way for the production of salidroside.
[0094] The present invention is disclosed above in the above-mentioned preferred embodiments, and is not a limitation on the implementation manners. For those of ordinary skill in the art, without departing from the concept of this patent, various different forms of changes and modifications can be made to the above-mentioned implementation manners, and these all fall within the protection scope of this patent. Therefore, the protection scope of this patent shall be subject to the claims.
Claims
1. A glycosyltransferase mutant, characterized in that, The mutant is based on the glycosyltransferase UGT shown in SEQ ID NO:1 BS in which isoleucine at position 62 is mutated to asparagine, serine at position 129 is mutated to threonine, phenylalanine at position 168 is mutated to tryptophan, and tyrosine at position 316 is mutated to serine simultaneously.
2. A gene encoding the glycosyltransferase mutant according to claim 1.
3. A recombinant vector carrying the gene according to claim 2.
4. A recombinant cell expressing the glycosyltransferase mutant according to claim 1, or containing the gene according to claim 2, or containing the recombinant vector according to claim 3.
5. The recombinant cell according to claim 4, wherein The recombinant cell uses bacteria or fungi as an expression host.
6. A method for catalytic synthesis of salidroside, characterized in that, The method is to add the glycosyltransferase mutant according to claim 1, or the recombinant cell according to claim 4 or 5, to a reaction system containing tyrosol, and react to prepare salidroside.
7. The method for catalytic synthesis of salidroside according to claim 6, characterized in that, The reaction system contains 1-10 mmol / L tyrosol, 2-20 mmol / L UDPG, 1-10% v / v DMSO, 5-40% v / v crude enzyme solution of the glycosyltransferase mutant, and 50-200 mmol / L Tris-HCl buffer; the enzyme concentration in the reaction system is 0.1-1 mg / ml.
8. Use of the glycosyltransferase mutant according to claim 1, or the gene according to claim 2, or the recombinant cell according to claim 4 or 5 in the catalytic synthesis of salidroside.
9. A catalyst, characterized in that, Containing the glycosyltransferase mutant according to claim 1, or the gene according to claim 2, or the recombinant cell according to claim 4 or 5.