CN110951799B

CN110951799B - Method for synthesizing (2S,3R) -p-methylsulfonyl phenyl serine by whole cell asymmetry of' one bacterium multienzyme

Info

Publication number: CN110951799B
Application number: CN201911340672.6A
Authority: CN
Inventors: 林娟; 许炼; 许鑫琦; 王力超; 赖凌燕; 陈承滔
Original assignee: Fujian Changsheng Biotechnology Development Co ltd; Fuzhou University
Current assignee: Fujian Changsheng Biotechnology Development Co.,Ltd.
Priority date: 2019-12-23
Filing date: 2019-12-23
Publication date: 2021-07-30
Anticipated expiration: 2039-12-23
Also published as: CN110951799A

Abstract

The invention provides a method for synthesizing (2S,3R) -p-methylsulfonylphenylserine in a whole cell asymmetric way by using one-bacterium multi-enzyme, belonging to the fields of biological catalysis and enzyme engineering. The asymmetric high-efficiency biosynthesis of (2S,3R) -p-methylsulfonylphenylserine is realized by simultaneously expressing L-threonine transaldolase (PsLTTA), alcohol dehydrogenase (ApADH) and formate dehydrogenase (CbFDH) in engineering bacteria of escherichia coli BL21(DE 3). The invention can obtain the key chiral building block (2S,3R) -p-methylsulfonylphenylserine of the beta-aminoalcohol antibiotic through one-step reaction at normal temperature and normal pressure, simultaneously removes the inhibition effect of byproduct acetaldehyde on PsLTTA, and is an efficient green biosynthesis way.

Description

Method for synthesizing (2S,3R) -p-methylsulfonyl phenyl serine by whole cell asymmetry of' one bacterium multienzyme

Technical Field

The invention belongs to the field of biological catalysis and enzyme engineering, and particularly relates to a method for synthesizing (2S,3R) -p-methylsulfonylphenylserine in a whole cell asymmetric manner by using 'one-bacterium multi-enzyme'.

Background

The (2S,3R) -p-methylsulfonylphenylserine is a key chiral building block for synthesizing beta-aminoalcohol antibiotics florfenicol and thiamphenicol. In 2015, only the export amount of the beta-aminoalcohol antibiotics in China exceeds 2700 tons, the year-by-year increase is 9.2 percent, and the export amount reaches 1.86 hundred million dollars, which is one of 10 major export drugs in China. Therefore, a rapid, efficient and green synthesis route of (2S,3R) -p-methylsulfonylphenylserine is established, and the method has a wide application prospect.

At present, the main method for producing (2S,3R) -p-methylsulfonyl phenyl serine in large scale at home and abroad takes p-methylsulfonyl benzaldehyde and glycine as raw materials, generates p-methylsulfonyl phenyl serine through aldol condensation reaction in the presence of copper sulfate and ammonia water, obtains p-methylsulfonyl phenyl serine ethyl ester through esterification, and obtains (2S,3R) -p-methylsulfonyl phenyl serine through D-tartaric acid resolution. Although the reaction route is simple and suitable for industrial production, the following problems exist: 1. the reaction conditions are severe, and strong acid and strong base are used simultaneously; 2. an expensive chiral resolving agent, D-tartaric acid, is required. Therefore, how to efficiently and asymmetrically synthesize the optically pure beta-hydroxy-alpha-amino acid in a green way is a hotspot and difficulty of the current chiral synthesis research.

Chemical synthesis route of (2S,3R) -p-methylsulfonylphenylserine ethyl ester in prior art

The prior art is provided with a patent number CN201710205793.4 provided by Kaiyuan national science and technology GmbH of Suzhou, the name: a synthetic method of p-methylsulfonyl phenyl serine ethyl ester is characterized in that diethyl sulfate and ethanol are added into a reaction kettle, and after the sulfuric acid and p-methylsulfonyl phenyl serine copper salt are subjected to reflux reaction, the reaction is finished by monitoring the esterification conversion rate to 95% -99%; and (3) cooling and crystallizing the mother liquor subjected to heat preservation and filter pressing, reducing the temperature to-5-0 ℃, performing suction filtration, performing water dissolution on a filter cake, removing copper from sodium sulfide, and performing ammonia precipitation on ammonia water to obtain the p-methylsulfonylphenylserine ethyl ester.

In addition, in the prior art, a gene recombinant plasmid of the chemical synthetase is firstly constructed, then the enzyme is produced by utilizing engineering bacteria for fermentation, and then the separation and purification of the enzyme are required. For example, in the synthesis of (2S,3R) -p-methylsulfonylphenylserine ethyl ester, a recombinant plasmid is constructed by a gene corresponding to a single protease, escherichia coli is transformed to express, and the protease is produced by fermentation of engineering bacteria, wherein the step is the fermentation production of the enzyme; then, the enzyme needs to be separated and purified to obtain pure enzyme, the operation is complicated and time-consuming, and the production cost is high. As disclosed in patent No. cn201810502755.x, title: a preparation method of D-p-methylsulfonylphenylserine ethyl ester takes DL-thermo-p-PMSE as a starting material, and the DL-threo-p-methylsulfonylphenylserine ethyl ester is resolved in a Tris-HCl buffer solution under the action of lipase (CAS NO: 9001-62-1). DL-thermo-p-PMSE is decomposed into D (-) -p-PMSE and L (+) -p-PMSE, the L (+) -p-PMSE is further hydrolyzed under the action of lipase without action on the D (-) -p-PMSE, so that the D (-) -p-MPSE can be directly obtained, and the highest ideal yield of the resolution is only 50%. The lipase described in this patent is obtained by separating lipase gene from bacterial strain, recombining it with vector plasmid, transforming Escherichia coli for heterologous expression, purifying the fermentation product by nickel column, and then using the purified lipase in the splitting process. The method has the advantages of complex and time-consuming operation and high production cost, and the method adopts a chiral resolution method instead of an asymmetric catalytic synthesis method, so that the yield of the target product is low and is only 50 percent.

Therefore, the prior art lacks a (2S,3R) -p-methylsulfonylphenylserine high-efficiency synthesis technology which is simple and convenient to operate, short in reaction time, free of the fermentation production and purification preparation process of enzyme and more beneficial to industrial production and application.

Disclosure of Invention

Aiming at the problems in the prior art, the invention provides a 'one-bacterium multi-enzyme' whole-cell catalysis method, which can obtain a key chiral building block (2S,3R) -p-methylsulfonylphenylserine of a beta-aminoalcohol antibiotic through one-step reaction at normal temperature and normal pressure, simultaneously removes the inhibition effect of byproduct acetaldehyde on L-threonine transaldolase PsLTTA, and is an efficient green biosynthesis way.

The specific invention content is as follows:

the method for synthesizing (2S,3R) -p-methylsulfonylphenylserine in a whole cell asymmetric way by using 'one bacterium multienzyme' is prepared by the following steps: step one, constructing a double-plasmid engineering bacterium

The L-threonine transaldolase is coded by a gene PsLTTA, and a recombinant expression plasmid pET28a-PsLTTA is constructed; the alcohol dehydrogenase and the formate dehydrogenase are respectively encoded by genes ApADH and CbFDH, and a recombinant expression plasmid pETDuet-ApADH/CbFDH is constructed; transforming the recombinant expression plasmids pETDuet-ApADH/CbFDH and pET28a-PsLTTA into escherichia coli BL21(DE3) to construct double-plasmid engineering bacteria; culturing the engineering bacteria to harvest thalli, and resuspending the thalli with Tris-HCl to obtain a whole-cell reaction solution of PsLTTA/ApADH/CbFDH.

Step two, synthesizing (2S,3R) -p-methylsulfonylphenylserine

Respectively putting 0.18-0.2 part of p-methylsulfonylbenzaldehyde, 0.18-0.2 part of L-threonine, 0.13-0.15 part of sodium formate, 0.035-0.04 part of reduced coenzyme I and 0.0013-0.0015 part of pyridoxal phosphate into a reaction container, and adding 10-12 parts of Tris-HCl buffer solution containing 10% ethyl acetate by volume into the reaction container in advance; fully and uniformly mixing, adding 0.25 part of wet whole cells, and carrying out oscillation reaction at 25-35 ℃ for 3.5-4.5 h; after the reaction was completed, the supernatant containing the product was collected.

Further, the synthesis principle is as follows: synthesizing (2S,3R) -p-methylsulfonylphenylserine and acetaldehyde by asymmetrically catalyzing p-methylsulfonylbenzaldehyde and L-threonine through L-threonine transaldolase, and removing the acetaldehyde generated in the catalytic reaction process of L-TTA through coupling of alcohol dehydrogenase and formate dehydrogenase so as to realize the cyclic regeneration of reduced coenzyme I in the catalytic process.

Further, the culture method of the engineering bacteria comprises the following steps: selecting monoclonal engineering bacteria cells containing recombinant plasmids, inoculating the monoclonal engineering bacteria cells into 20mL LB liquid culture medium containing 100 ug/mL ampicillin and 50 ug/mL kanamycin, and culturing at 37 ℃ and 180rpm overnight; sucking 10mL of culture solution, transferring the culture solution into 1L of LB liquid culture medium, and carrying out shaking culture at 37 ℃ and 200 rpm; when OD is reached₆₀₀When the concentration is 0.5, adding inducer for inducing the expression of L-threonine transaldolase, alcohol dehydrogenase and formate dehydrogenase proteins, adding inducer to the mixture to a final concentration of 0.1mM, inducing the mixture at 28 ℃ for 16h, stopping culturing, and centrifuging the mixture at 5000rpm and 4 ℃ for 10min to obtain thalli.

Further, the inducer is isopropyl-beta-D-thiogalactoside.

Further, the cells described in step one were resuspended in Tris-HCl at a concentration of 100mM and pH 7.0 to prepare a 250mg/mL stock solution as the whole cell reaction solution.

The invention has the following beneficial effects: 1. simultaneously expressing L-threonine transaldolase (PsLTTA), alcohol dehydrogenase (ApADH) and formate dehydrogenase (CbFDH) in engineering bacteria of escherichia coli BL21(DE3), and realizing high-efficiency synthesis of (2S,3R) -p-methylsulfonylphenylserine by using a whole-cell catalysis method; the PsLTTA can asymmetrically catalyze p-methylsulfonylbenzaldehyde and L-threonine to synthesize (2S,3R) -p-methylsulfonylphenylserine and acetaldehyde; ApADH can catalyze acetaldehyde to be converted into ethanol, and the inhibition effect of acetaldehyde on PsLTTA is relieved; CbFDH can realize NADH/NAD in system⁺And (4) cyclic regeneration. The method can obtain the key chiral building block (2S,3R) -p-methylsulfonylphenylserine of the beta-aminoalcohol antibiotic through one-step reaction at normal temperature and normal pressure, simultaneously removes the inhibition effect of byproduct acetaldehyde on PsLTTA, and is an efficient green biosynthesis way; in addition, the conversion rate of the substrate can reach 99% after the method reacts for 4 hours at the temperature of 30 ℃, and the enantiomeric excess ee of the product can reach ee>99.9%, diastereomer excess de 94.1%; compared with the yield of the product (2S,3R) -p-methylsulfonylphenylserine which is catalyzed by only adopting PsLTTA, the yield of the product is improved by 4.3 times.

2. Compared with the method for carrying out catalytic reaction by using a threonine transaldolase, alcohol dehydrogenase and glucose dehydrogenase pure enzyme preparation, the one-bacterium multi-enzyme whole-cell catalytic method has the advantages of simple and convenient operation and short reaction time, omits the fermentation production and purification preparation process of the enzyme, reduces the production cost and is more beneficial to industrial production and application.

Drawings

FIG. 1 shows the chemical principle of the reaction for synthesizing (2S,3R) -p-methylsulfonylphenylserine according to the present invention;

FIG. 2 is an SDS-PAGE electrophoresis chart of the crushed supernatant of PsLTTA/ApADH/CbFDH BL21 engineering bacteria;

FIG. 3 is a HPLC check chart of (2S,3R) -p-methylsulfonylphenylserine.

Detailed Description

The following preferred embodiments of the present invention are provided to aid in a further understanding of the invention. It should be understood by those skilled in the art that the description of the embodiments of the present invention is by way of example only, and not by way of limitation.

The L-threonine transaldolase is coded by a gene PsLTTA, and a recombinant expression plasmid pET28a-PsLTTA is constructed; the alcohol dehydrogenase and the formate dehydrogenase are respectively encoded by genes ApADH and CbFDH, and a recombinant expression plasmid pETDuet-ApADH/CbFDH is constructed; transforming the recombinant expression plasmids pETDuet-ApADH/CbFDH and pET28a-PsLTTA into escherichia coli BL21(DE3) to construct double-plasmid engineering bacteria; culturing the engineering bacteria to harvest thalli, resuspending the thalli with Tris-HCl with the concentration of 100mM and the pH value of 7.0, and preparing into 250mg/mL mother solution to obtain a whole-cell reaction solution of PsLTTA/ApADH/CbFDH; step two, synthesizing (2S,3R) -p-methylsulfonylphenylserine

As shown in FIG. 1, the synthesis principle of the invention is as follows: synthesizing (2S,3R) -p-methylsulfonylphenylserine and acetaldehyde by asymmetrically catalyzing p-methylsulfonylbenzaldehyde and L-threonine through L-threonine transaldolase, and removing the acetaldehyde generated in the catalytic reaction process of L-TTA through coupling of alcohol dehydrogenase and formate dehydrogenase so as to realize the cyclic regeneration of reduced coenzyme I in the catalytic process.

Further, the culture method of the engineering bacteria comprises the following steps: selecting monoclonal engineering bacteria cells containing recombinant plasmids, inoculating the monoclonal engineering bacteria cells into 20mL LB liquid culture medium containing 100 ug/mL ampicillin and 50 ug/mL kanamycin, and culturing at 37 ℃ and 180rpm overnight; sucking 10mL of culture solution, transferring the culture solution into 1L of LB liquid culture medium, and carrying out shaking culture at 37 ℃ and 200 rpm; when OD is reached₆₀₀When the content of the organic acid is 0.5,adding inducer for inducing the expression of L-threonine transaldolase, alcohol dehydrogenase and formate dehydrogenase proteins, adding inducer isopropyl-beta-D-thiogalactoside with final concentration of 0.1mM, inducing at 28 deg.C for 16h, stopping culturing, centrifuging at 4 deg.C and 5000rpm for 10min, and harvesting thallus.

The inventors specifically set up the following experiments to verify the present invention.

Test 1

Fermentation preparation for constructing whole-cell reaction liquid of two-plasmid engineering bacteria PsLTTA/ApADH/CbFDH

L-threonine transaldolase (PsLTTA, the amino acid sequence of which is shown in SEQ ID NO: 1) is encoded by gene PsLTTA (the base sequence of which is shown in SEQ ID NO: 4), and recombinant expression plasmid pET28a-PsLTTA is constructed. Alcohol dehydrogenase (ApADH, amino acid sequence shown in SEQ ID NO: 2) and formate dehydrogenase (CbFDH, amino acid sequence shown in SEQ ID NO: 3), encoded by the genes ApADH (base sequence shown in SEQ ID NO: 5) and CbFDH (base sequence shown in SEQ ID NO: 6), respectively, to construct a recombinant expression plasmid pETDuet-ApADH/CbFDH. The recombinant expression plasmids pETDuet-ApADH/CbFDH and pET28a-PsLTTA are transformed into Escherichia coli BL21(DE3) to construct the double-plasmid engineering bacteria. A single clone containing the recombinant plasmid was picked up and inoculated into 20mL of LB liquid medium (containing 100. mu.g/mL of ampicillin and 50. mu.g/mL of kanamycin), and cultured overnight at 37 ℃ and 180 rpm. 10mL of the culture medium was pipetted into 1L of LB liquid medium (containing 100. mu.g/mL ampicillin and 50. mu.g/mL kanamycin), and cultured at 37 ℃ with shaking at 200 rpm. When OD is reached₆₀₀When the concentration is 0.5, IPTG is added to the mixture to a final concentration of 0.1mM, the culture is stopped after 16 hours of induction at 28 ℃, and the mixture is centrifuged at 5000rpm for 10 minutes at 4 ℃ to obtain the thalli.

In addition, a single variable contrast group which is not added with IPTG induction is arranged to obtain thalli. The results shown in FIG. 2 were obtained by SDS-PAGE detection, and lane 1 is a control group not induced by IPTG;

lanes

2 and 3 are induced by adding 100Mm IPTG, and specific bands of L-threonine transaldolase, alcohol dehydrogenase and formate dehydrogenase appear near 48kDa, 40kDa and 45kDa, respectively, as shown in fig. 2, indicating that the 3 proteins are successfully expressed, verifying the feasibility of the method of the present invention to utilize whole cell reaction liquid for fermentation.

Test 2

(1) Asymmetric catalytic synthesis of (2S,3R) -p-methylsulfonylphenylserine by using' one bacterium multi-enzyme

10mL of reaction system: 0.18g of p-methylsulfonylbenzaldehyde, 0.18g L-threonine, 0.13g of sodium formate, 35mg of NADH, and 1.3mg of PLP were put into a 50mL reaction flask, to which 10mL of Tris-HCl buffer (100mM, pH7) containing 10% ethyl acetate (v/v) was added. After mixing well, 0.25g of whole cells (wet weight) was added and reacted at 30 ℃ for 4 hours with shaking. After the reaction is finished, centrifuging at 10000rpm for 5min to remove cell precipitates, collecting supernatant containing products, adding methanol according to a ratio of 1:20, storing at 4 ℃, and taking a whole cell reaction only expressing PsLTTA as a control.

(2) OPA/NAC derivatization detection product (2S,3R) -p-methylsulfonylphenylserine

OPA/NAC derivatization reagent preparation: solution A: weighing 10mg of o-phthalaldehyde (OPA) and adding the o-phthalaldehyde (OPA) into 5mL of methanol, and oscillating the mixture at 30 ℃ to fully dissolve the o-phthalaldehyde; and B, liquid B: 10mg of N-acetylcysteine (NAC) was weighed out and dissolved in 20mL of buffer (0.2M boric acid, 0.2M potassium chloride); mixing solution A and solution B, and storing at 4 deg.C in dark.

And (3) uniformly mixing 100 mu L of reaction supernatant and 400 mu L of OPA/NAC derivatization reagent, standing at room temperature in the dark for 10min, and detecting by High Performance Liquid Chromatography (HPLC). The HPLC detection conditions are as follows: detection wavelength: 236 and 340 nm; a chromatographic column: agilent C18 column (250X 4.6mm,5 μm); mobile phase: 50mM KH₂PO₄pH 7.0/acetonitrile (79/21, v/v); flow rate: 1mL/min, temperature: 30 ℃; loading: 20 μ L.

The HPLC detection result is shown in figure 3, wherein, the A picture adopts PsLTTA/ApADH/CbFDH whole cell catalysis; panel B was catalyzed with PsLTTA whole cells. T is_(2S,3R)＝5.7min，T_(2S,3S)6.5 min. As shown in fig. 3, the peak-off times of the products (2S,3R) -p-methylsulfonylphenylserine and (2S, 3S) -p-methylsulfonylphenylserine were 5.7min and 6.5min, respectively; by calculating and comparing the peak areas of HPLC, the enantiomeric excess ee of the product can be obtained>99.9%, diastereomer excess de 94.1%, and 99% substrate conversion. Compared with the yield of the product (2S,3R) -p-methylsulfonylphenylserine which is catalyzed by only adopting PsLTTA, the yield of the product is improved by 4.3 times. Therefore, the method of the invention is more suitable for preparationSimple, fast and high yield.

It will be evident to those skilled in the art that the invention is not limited to the details of the foregoing illustrative embodiments, and that the present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof. The present embodiments are therefore to be considered in all respects as illustrative and not restrictive, the scope of the invention being indicated by the appended claims rather than by the foregoing description, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein.

Sequence listing

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Fujian Changsheng Biotechnology Development Co.,Ltd.

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<400> 5

atgagcggta aaatgaaagc agccgttgtt catgaatttg gtaaaccgct gaccattgag 60

gaactggata ttccgcctat taaaccgaca cagattctgg ttaaaatgat tgcctgtggt 120

gtttgtcata ccgatctgca tgccgcaagc ggtgattggc cgaaaaaacc gcatctgccg 180

tttattccgg gtcatgaagg tgttggtaca gttgttcagg ttggtagcga agttgattgg 240

gttaaagaag gtgacgttgt tggcgttccg tggctgtata gcgcatgtgg tcattgtgaa 300

cattgtctgg caggttggga aaccctgtgt gcaaaacaag aagaaaccgg ttatagcgtg 360

aatggttgtt ttgccgaata tgttgttgca gacccgaact atattgcaca tctgccgaaa 420

ggtgttgatc cggttaaagt tgcaccggtt ctgtgtgcag gtctgaccgt gtataaaggt 480

ctgaaaatga ccgatacacg tgcaggtaat tgggttgcaa ttagcggtgt tggtggtctg 540

ggtcagatgg cagttcagta tgcagttgca atgggtctga atgttgtggc agttgatatc 600

gatgatgaaa aactggcaac cgcaaaaaaa ctgggtgcaa cctataccgt taatgcacgt 660

aataccgatc cggcagcatt tatgcaagaa aaagttggtg gtgttcatgg tggcctgatt 720

accgcagtta gcaccaaagc atttagccag gcaatgggtt atgttcgtgc cggtggcacc 780

ctggttctga atggtctgcc tccgggtgat tttccgatta gcatttttga tatggtgatg 840

aacgccatta ccattcgtgg tagcattgtt ggcacccgtc tggatatgat tgaagcactg 900

agcttttttg cagaaggtaa agttaccagc gttaccacca ccgatcgtat tgataacatt 960

aacgcaattt tcgacgccct gaaaaatggt cgtgttgaag gtcgtgtggt tctggatttt 1020

cgcaattaa 1029

<210> 6

<211> 1095

<212> DNA

<213> Artificial sequence ()

<400> 6

atgaaaattg tgctggtgct gtatgatgca ggtaaacatg cagcagatga agaaaaactg 60

tatggctgca ccgaaaataa actgggtatt gcaaattggc tgaaagatca gggtcatgaa 120

ctgattacca ccagtgataa agaaggtgaa accagcgaac tggataaaca tattccggat 180

gccgatatta tcattaccac accgtttcat ccggcatata tcaccaaaga acgtctggat 240

aaagccaaaa atctgaaact ggttgttgtt gccggtgttg gtagcgatca tattgatctg 300

gattatatca atcagaccgg caaaaaaatc agcgttctgg aagttaccgg tagcaatgtt 360

gttagcgttg cagaacatgt tgttatgacc atgctggttc tggttcgcaa ttttgttccg 420

gcacatgagc agattattaa ccatgattgg gaagttgcag ccattgcaaa agatgcctat 480

gatattgaag gtaaaaccat tgcaaccatt ggtgcaggtc gtattggtta tcgtgttctg 540

gaacgtctgc tgccgtttaa tccgaaagaa ctgctgtatt atgattatca ggcactgccg 600

aaagaagccg aagaaaaagt tggtgcccgt cgtgttgaaa atattgaaga actggttgca 660

caggccgata ttgttaccgt taatgcaccg ctgcatgccg gtacaaaagg tctgattaac 720

aaagagctgc tgagcaaatt caaaaaaggt gcatggctgg ttaataccgc acgtggtgca 780

atttgtgttg ccgaagatgt tgcagcagca ctggaaagcg gtcagctgcg tggttatggt 840

ggtgatgttt ggtttccgca gccagcaccg aaagatcatc cgtggcgtga tatgcgtaac 900

aaatatggtg ccggtaatgc aatgacaccg cattatagcg gtacaaccct ggatgcacag 960

acccgttatg cagaaggcac caaaaacatt ctggaaagct ttttcaccgg caaatttgat 1020

tatcgtccgc aggatattat tctgctgaat ggtgaatatg tgaccaaagc ctatggcaaa 1080

cacgacaaaa aataa 1095

Claims

A method for the whole-cell asymmetric synthesis of (2S,3R) -p-methylsulfonylphenylserine by using one bacterium and multiple enzymes is characterized by comprising the following steps:

step one, constructing a double-plasmid engineering bacterium

L-threonine transaldolase, PsLTTA, the amino acid sequence of which is set forth in SEQ ID NO: 1, is composed of a genePsLTTAAnd the base sequence is shown as SEQ ID NO: 4, constructing a recombinant expression plasmid pET28 a-PsLTTA;

alcohol dehydrogenase, ApADH, amino acid sequence as shown in SEQ ID NO: 2, the amino acid sequence of formate dehydrogenase, CbFDH is shown as SEQ ID NO: 3, each consisting of a geneApADHAnd the base sequence is shown as SEQ ID NO: 5 is shown in andCbFDHthe base sequence is shown as SEQ ID NO: 6, constructing a recombinant expression plasmid pETDuet-ApADH/CbFDH; the recombinant expression plasmids pETDuet-ApADH/CbFDH and pET28a-PsLTTA are transformed into Escherichia coli BL21(DE3) constructing double-plasmid engineering bacteria;

culturing the engineering bacteria to harvest thalli, and resuspending the thalli with Tris-HCl to obtain a whole-cell reaction solution of PsLTTA/ApADH/CbFDH;

step two, synthesizing (2S,3R) -p-methylsulfonylphenylserine, respectively taking 0.18-0.2 part of p-methylsulfonylbenzaldehyde, 0.18-0.2 part of L-threonine, 0.13-0.15 part of sodium formate, 0.035-0.04 part of reduced coenzyme I and 0.0013-0.0015 part of pyridoxal phosphate by mass, putting into a reaction container, and adding 10-12 parts of Tris-HCl buffer solution containing 10% ethyl acetate by volume into the reaction container in advance, wherein the pH value of the Tris-HCl buffer solution is 7.0; after fully and uniformly mixing, adding whole cells with the wet weight of 0.25 part, and carrying out oscillation reaction for 4 hours at the temperature of 30 ℃; after the reaction is finished, collecting the supernatant containing the product;

the synthesis principle is as follows: synthesizing (2S,3R) -p-methylsulfonylphenylserine and acetaldehyde by asymmetrically catalyzing p-methylsulfonylbenzaldehyde and L-threonine through L-threonine transaldolase, and removing the acetaldehyde generated in the catalytic reaction process of the L-threonine transaldolase through coupling of alcohol dehydrogenase and formate dehydrogenase and realizing the cyclic regeneration of reduced coenzyme I in the catalytic process.
2. The method for the whole-cell asymmetric synthesis of (2S,3R) -p-methylsulfonylphenylserine according to claim 1, which comprises the following steps: the culture method of the engineering bacteria comprises the following steps: selecting monoclonal engineering bacteria cells containing recombinant plasmids, inoculating the monoclonal engineering bacteria cells into 20mL LB liquid culture medium containing 100 ug/mL ampicillin and 50 ug/mL kanamycin, and culturing at 37 ℃ and 180rpm overnight; sucking 10mL of culture solution, transferring the culture solution into 1L of LB liquid culture medium, and carrying out shaking culture at 37 ℃ and 200 rpm; when OD600 is 0.5, adding inducer for inducing the expression of L-threonine transaldolase, alcohol dehydrogenase and formate dehydrogenase proteins, adding inducer with final concentration of 0.1mM, inducing at 28 deg.C for 16h, stopping culturing, centrifuging at 4 deg.C and 5000rpm for 10min, and harvesting thallus; the inducer is isopropyl-beta-D-thiogalactoside.
3. The method for the whole-cell asymmetric synthesis of (2S,3R) -p-methylsulfonylphenylserine according to claim 1, which comprises the following steps: and step one, the thalli is resuspended by using Tris-HCl with the concentration of 100mM, the pH value is 7.0, and a mother solution of 250mg/mL is prepared and used as the whole cell reaction solution.