CN114875005A

CN114875005A - Construction and application of enantioselectively inverted omega-transaminase mutant

Info

Publication number: CN114875005A
Application number: CN202110162624.3A
Authority: CN
Inventors: 林双君; 刘琦; 邓子新; 黄婷婷
Original assignee: Shanghai Jiaotong University
Current assignee: Shanghai Jiaotong University
Priority date: 2021-02-05
Filing date: 2021-02-05
Publication date: 2022-08-09
Anticipated expiration: 2041-02-05
Also published as: CN114875005B

Abstract

The invention provides an enantioselectively inverted omega-transaminase ATA117 mutant, and the protein amino acid sequence of the mutant is shown as SEQ ID No. 1; the second aspect of the invention provides a nucleotide sequence for coding the omega-transaminase ATA117 mutant, which is shown as SEQ ID No. 2; in a third aspect of the invention, there is provided a plasmid carrying the above nucleotide sequence; in the fourth aspect of the invention, the invention provides a genetically engineered bacterium for expressing the omega-transaminase ATA117 mutant; in the fifth aspect of the invention, a method for producing (1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylpropane 2b by using an omega-transaminase ATA117 mutant is provided. The enantiomer ratio of the ATA117_ ACHH mutant provided by the invention is 11.7E (R), the enantiomer ratio (E (S) ═ 8.5) of the wild ATA117 to the chiral 1, 3-dihydroxy-1-p-methylsulfonyl phenyl acetone 1 is reversed, and the mutant provided by the invention lays a technical process foundation for the application of the mutant in the synthesis of chiral (1R,2R) -aminodiol products.

Description

Construction and application of enantioselectively inverted omega-transaminase mutant

Technical Field

The invention relates to the field of enzyme engineering, in particular to construction and application of an enantioselectively inverted omega-transaminase mutant.

Background

Transaminases are divided into two classes, alpha-and omega-transaminases. Alpha-aminotransferases include subfamilies I, III and IV, which catalyze the transamination of alpha-carboxylic acids that depend on the substrate ketone. Omega-aminotransferases belong to the type II subfamily, are alpha-carboxylic acids independent of substrate ketones, and are widely applied to asymmetric transamination of various aliphatic ketones and aromatic ketones to generate stereospecific amines. Among them, ATA117 is one of the widely used ω -transaminases. Wolfgang Kroutil project group reported in 2008 that ATA117 can efficiently catalyze a series of aromatic ketones such as 4-phenyl-2-butanone and aliphatic ketones such as octanone to produce amines with R configuration, and ee is more than or equal to 99%. The group of Wolfgang Kroutil subjects reported in 2010 that ATA117 catalyzed organoselenium acetophenone to produce stereospecific R-para-selenophenylethylamine, a chiral ligand for palladium catalysts. In the same year, the Gregory J.Hughes subject group obtains an ATA-117-Rd11 mutant by carrying out multiple rounds of enzyme evolution on ATA117, and the mutant realizes effective catalysis on a large steric hindrance substrate sitagliptin precursor ketone. While the Yu-Guo Zheng project group catalyzed the generation of stereospecific L-glufosinate product with ATA-117-Rd11 mutant in 2020, ee > 99%, 80kg of the reaction precursor was converted to 70kg of L-glufosinate product in 24 hours. The results show that the substrate spectrum of the omega-transaminase ATA117 is wide, and the omega-transaminase ATA117 can efficiently catalyze various substrates to generate stereospecific amine.

In addition to producing stereospecific amines, ω -aminotransferases also have a certain enantioselectivity towards ortho chiral centers. The group of Wolfgang krotil topic reported in 2009 that R-4-phenyl-2-pyrrolidone was obtained by kinetic resolution of racemic aldehyde substrates by the ω -transaminase ATA 117. ATA117 has enantioselectivity to the chiral center at the ortho position of aldehyde, and the ee of the transaminated product can reach 68% through the optimization of cosolvent and pH. The kinetic resolution of racemic α -alkyl- β -ketoesters by 24 commercial S-and R-transaminases was examined by the Vicente Gotor project group in 2013, and the results showed that the catalytic efficiency was high, but the diastereoselectivity was poor. The kinetic resolution of racemic 2-phenylpropionaldehyde and para-meta-ortho methyl and methoxy substituted 2-phenylpropionaldehyde by Pseudomonas, Arthrobacter, Aspergillus and Sinomonas omega-aminotransferases was investigated in 2014 by the Wolfgang Krotil group, and the stereoselectivity of these aminotransferases to the ortho chiral center was characterized, with ee (R) 76-98%. In 2018, the Wolfgang Kroutil subject group investigates the kinetic resolution condition of omega-aminotransferase from Arthrobacter KNK168, Aspergillus terreus, Fusarium graminearum, hyphomomonas, New Saccharomycotina, Bacillus megaterium and the like on racemic fatty aldehyde, and obtains the antiepileptic drugs of Buvalsartan and Pregabalin with high stereoselectivity through one-step transamination reaction. Wherein, the omega-aminotransferase from the raw silk monad can obtain the R-brivaracetam chiral intermediate with high stereoselectivity through one-step kinetic resolution, and the ee can reach 92 percent. The Wolfgang Kroutil subject group respectively makes mutation of a plurality of amino acid sites on the basis of an omega-transaminase and an ArRmut11 mutant from fusarium graminearum, the obtained mutant catalyzes racemic fatty aldehyde to synthesize S-pregabalin, and ee of the S-pregabalin can respectively reach 76% and 80%. In 2019, the group of the Iv & ltn Lavandera issues reports the kinetic resolution of a series of commercial aminotransferases and aminotransferases from bacillus megaterium and mutants thereof on racemic alpha-alkyl-beta-ketoamide substrates, wherein the commercial R aminotransferase has the highest catalytic efficiency and stereoselectivity on the substrates, and the diastereoselectivity de can reach 96%. The transaminase derived from the bacillus megaterium and the mutant thereof can convert only part of substrates, and has low diastereoselectivity. In summary, the literature reports mostly show that high stereoselectivity for the chiral center in ortho position to aldehyde substrates can be obtained by screening omega-transaminases from different species, whereas high stereoselectivity for the chiral center in ortho position to ketone substrates such as α -alkyl- β -ketoamides can be obtained by screening commercial transaminases. However, there is no report in the literature on the enantioselectivity of ω -transaminase to chiral aromatic hydroxyketone compounds.

Therefore, it is a problem to be solved by those skilled in the art how to provide a ω -transaminase with high enantioselectivity to chiral aromatic hydroxy ketone compounds, so that the ω -transaminase can be applied to the synthesis of (1R,2R) -aminodiol products by kinetic resolution.

Disclosure of Invention

At present, how to provide a omega-aminotransferase with high enantioselectivity to chiral aromatic hydroxyketone compounds, which is applied to the kinetic resolution synthesis of (1R,2R) -aminodiol products, is a problem to be solved urgently for those skilled in the art.

In order to solve the technical problems, the invention carries out molecular modification on omega-transaminase ATA117 based on semi-rational design, and combines an iterative saturation mutation method of genes to obtain corresponding mutants.

The invention provides an enantioselectively inverted omega-transaminase ATA117 mutant, and the protein amino acid sequence of the mutant is shown as SEQ ID No. 1.

Further, the mutant is produced by iterative saturation mutation of the amino acid sequence of ATA117, and has mutation of at least one of valine at position N1, phenylalanine at position N2, isoleucine at position N3, and phenylalanine at position N4.

Further, valine at position N1 was mutated to alanine.

Further, phenylalanine at position N2 was mutated to cysteine.

Further, isoleucine at position N3 was mutated to histidine.

Further, phenylalanine at position N4 was mutated to histidine.

Furthermore, the mutant has higher R-enantioselectivity to racemic p-methylsulfonyl phenyl dihydroxy ketone 1.

In a second aspect of the invention, there is provided a DNA comprising a nucleotide sequence encoding the ω -transaminase ATA117 mutant.

Further, the nucleotide sequence of the DNA is shown in SEQ ID No. 2.

In a third aspect of the invention, there is provided a plasmid carrying the above nucleotide sequence.

Furthermore, the recombinant plasmid carrying the gene encoding omega-transaminase ATA117 was obtained by inserting the ATA117 gene synthesized by Hongxn Biotechnology Ltd into pRSFduet-1 vector sal I/Hind III cleavage site by digestion ligation.

In the fourth aspect of the invention, the invention provides a genetically engineered bacterium for expressing the omega-transaminase ATA117 mutant.

Further, the construction method of the genetic engineering bacteria comprises the following steps: a1, using recombinant plasmid carrying gene coding ATA117 as template, A2, designing and synthesizing primer containing mutation, A3, reverse PCR amplifying whole plasmid, A4, PCR product transforming expression host.

Further, the expression host is E.coli BL21(DE3)

In the fifth aspect of the invention, a method for producing (1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylpropane 2b by using an omega-transaminase ATA117 mutant is provided: the method takes omega-transaminase ATA117 mutant pure protein as a catalyst, catalyzes racemic 1, 3-dihydroxy-1-p-methylsulfonyl propiophenone 1 in a buffer system, and obtains (1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonyl phenylpropane 2b with high stereoselectivity through kinetic resolution.

Further, the buffer system is a single aqueous phase system.

The invention has the following effects:

1. the ATA117_ ACHH mutant obtained by the invention has higher R-selectivity on the ortho chiral center of the aromatic hydroxyketone compound, and supplements the research on the enantioselectivity of omega-aminotransferase on the chiral hydroxyketone compound.

2. The kinetic resolution of the mutant ATA117_ ACHH to the racemic hydroxyketone compound can obtain the (1R,2R) -amino alcohol with high stereoselectivity, and the (1R,2R) -amino alcohol is a chiral intermediate of various drug intermediates, such as thiamphenicol, florfenicol and pseudoephedrine, so the mutant has the potential of being applied to the synthesis of chiral drug intermediates.

3. The enantiomer ratio of the wild ATA117 to the chiral 1, 3-dihydroxy-1-p-methylsulfonylphenylacetone 1 is 8.5E (S), and the enantiomer ratio of the ATA117_ ACHH mutant is 11.7E (R), so that the mutant obtained by the invention lays a foundation for the application of the mutant in the synthesis of chiral (1R,2R) -aminodiol products.

Drawings

FIG. 1, schematic representation of the kinetic resolution of ATA117_ ACHH mutant for Compound 1;

FIG. 2, graph showing the conversion rate of ATA117_ ACHH mutant protein kinetic resolution racemic substrate 1 synthesis 2;

FIG. 3, stereoselective scheme for kinetic resolution of protein from ATA117_ ACHH mutant racemic substrate 1 Synthesis 2.

Detailed Description

The present invention is described in further detail below with reference to specific examples and with reference to the data. It will be understood that these examples are intended to illustrate the invention and are not intended to limit the scope of the invention in any way.

General description of the sources of the biological materials described in the present invention:

(1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylpropane was obtained from Huang Shi Yongxin Biotech, Inc. A common liquid chromatographic column C18(4.6mm multiplied by 250mm, particle size 5 μm) is a product of Philomena scientific instruments company; the chiral liquid chromatographic column IG (4.6mm × 250mm, particle size 5 μm) is a product of Daiiluo pharmaceutical chiral technology company.

Example 1 iterative saturation mutagenesis

VN-F：agcgacgttacctataccnnkttccacgtttggaacggtaacgca

VN-R：accgttccaaacgtggaamnnggtataggtaacgtcgctatgcag

FN-F：accgaactgcgcgaagcgnnkgttagcgttagcattacccgcggt

FN-R：ggtaatgctaacgctaacmnncgcttcgcgcagttcggtttttgc

IN-F:gctgttccgtaccagtggnnkgttccgtttgatcgtattcgcgac

IN-R:aatacgatcaaacggaacmnnccactggtacggaacagcgtacat

FN-F:ctggcagaaggtagcggcnnkaacgtcgtcgtcattaaagatggg

FN-R:tttaatgacgacgacgttmnngccgctaccttctgccagtaaacc

Wherein N represents A, T, C and G arbitrary bases, K represents T or G bases, and the degenerate codon can code 20 random amino acids.

(1) PCR amplification (95 ℃ for 5 min; 95 ℃ for 30s, 65 ℃ for 30s, 72 ℃ for 5min, 34 cycles; 72 ℃ for 10min) with the recombinant plasmid pRSFduet-1-ATA117 as a template and VN-F and VN-R as primers; the PCR product is directly transformed into E.coli BL21(DE3) competent cells after being digested by FD-Dpn I (30 ℃ incubator, 6h), the recovery solution is fully blown and evenly mixed, about 1/8 recovery solution is coated on a kanamycin-resistant LB plate, and the kanamycin-resistant LB plate is cultured for 12-16h at 37 ℃ to obtain a recombinant sub-library.

(2) All single colonies (about 40) on the dilution plate were picked and cultured in kanamycin-resistant LB medium at 37 ℃ for 7-8h, a portion of the culture was used for sequencing, and the other portion of the culture was temporarily stored in a refrigerator at 4 ℃ for a short period of time. Transferring the positive transformant culture solution to a fresh 24-hole deep-hole plate filled with 5mL LB culture medium containing kanamycin resistance by 1 percent of inoculation amount, culturing for 3h at 37 ℃, adding 0.2mM IPTG inducer, culturing for 15h at 20 ℃, inducing the high-efficiency expression of recombinant genes, centrifugally collecting thalli from recombinant cells at 4000rpm, and preserving at-80 ℃. pRSFduet-1-ATA117 recombinant cells were obtained by the same induction method as a control.

(3) The obtained recombinant cells were suspended in 300ul of 50mM Tris-Cl (pH 7.5), sonicated, centrifuged at low temperature, and the supernatant was used for the catalytic reaction of racemic 1, 3-dihydroxy-1-p-methylsulfonylpropiophenone.

(4) The reaction product of (3) was subjected to a C18 column to determine its diastereoselectivity, and the conditions of liquid phase analysis were as described in example 2. The experimental result shows that in the ATA117/VN saturation mutant, the selectivity of the ATA117_ A mutant on the S-hydroxyketone substrate is obviously reduced compared with that of the wild type ATA117, so that ATA117_ A is selected as a template for the next round of FN saturation mutation. The mutant construction, expression, reaction and detection methods are as described above. The experimental result shows that IN the ATA117_ A/FN saturated mutant, the selectivity of the ATA117_ AC mutant to the chiral hydroxyketone substrate is reversed, so that the ATA117_ AC is selected as a template of the next round of IN saturated mutation. After IN saturation mutation, FN mutation is iterated, and finally the mutant ATA117_ ACHH with higher selectivity on R-hydroxyketone substrate is obtained.

Example 2 purification of transaminase expression

Expression and purification method of transaminase: coli BL21(DE3) single colonies containing the recombinant plasmid were picked up on LB solid medium, inoculated into 40ml of LB liquid medium (containing 50. mu.g/ml kanamycin antibiotic), and cultured overnight at 37 ℃ and 220 rpm. Transferring 7.5ml of the bacterial culture solution to a 2L shake flask containing 500ml of liquid LB medium, inoculating two flasks, culturing at 37 ℃ and 220rpm until OD600 reaches 0.6-0.8, adding 0.2mM IPTG for induction, and performing induction culture at 20 ℃ and 200rpm for 15 h. 1L of the fermentation broth was collected, centrifuged at 5000rpm for 20min to collect the cells, and the collected cells were resuspended in 30mL of nickel column binding buffer and placed in an ice-water mixture. Ultrasonic crushing conditions: the operation is carried out for 5s and the pause is carried out for 10s for 30 min. The mixture after disruption treatment was centrifuged at 12,000rpm for 1 hour, and the supernatant was filtered through a 0.22 μm filter. The filtered sample was loaded with 2ml of nickel packing pre-equilibrated with nickel column binding buffer, the hetero-proteins were washed with 10 column volumes of 50mM imidazole elution buffer, and then the target proteins were eluted with 5ml of 250mM imidazole elution buffer, and the samples were pipetted, 500ul per tube. And (3) measuring the protein concentration of each tube by using the NanoDrop, combining several tubes of protein liquid with higher concentration, diluting or concentrating to 2.5ml, loading a desalting column balanced by glycerol buffer solution, draining the protein liquid, and adding 3.5ml of glycerol buffer solution to elute the protein.

Example 3 determination of the enantioselectivity of ATA117 and its mutants

The reaction system consisted of 100ul (50mM Tris-Cl buffer, pH 7.5), 2mM racemic 1, 3-dihydroxy-1-p-methylsulfonylpropiophenone, 2mM PLP, 200mM D-Ala, 5uM ATA117, 20uM ATA117_ A, 20uM ATA117_ AC, 40uM ATA117_ ACH, 40uM ATA117_ ACHH, and was reacted at 30 ℃ for 1h, and the reaction was terminated by adding 200ul methanol after the reaction was completed. The sample analysis employed an Agilent liquid phase, C18 column (4.6 mm. times.150 mm, particle size 3 μm), column temperature 30 ℃, 224nm, 0.5ml/min, phase A: h ₂ O(10mM KH ₂ PO ₄ pH 8.5), phase B: acetonitrile, chromatography conditions as shown in table 1:

TABLE 1 chromatography Condition Table

The retention time of (1S,2R) -and (1R,2R) -1, 3-dihydroxy-1-p-methylsulfonylpropiophenone is 13.2 min and 14.1min respectively. de ═ (RR-SR)/(RR + SR), E ═ ln [1-c × (1+ de) ]/ln [1-c × (1-de) ]. Wherein: RR and SR represent the concentrations of (1R,2R) -and (1S,2R) -1, 3-dihydroxy-1-p-methylsulfonylpropylphenyl ketone, and c represents the conversion rate of racemic 1, 3-dihydroxy-1-p-methylsulfonylpropylphenyl ketone.

The results are shown in table 2:

TABLE 2 enantiomeric ratio (E) of ATA117 and mutants thereof

De represents the diastereomeric excess value

Example 4 Synthesis of (1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylpropane using ATA117_ ACHH reaction System total 100ul (100mM Tris-Cl buffer, pH 7.5), 25uM ATA117_ ACHH pure protein, 5mM racemic 1, 3-dihydroxy-1-p-methylsulfonylphenylacetone, 2mM PLP, 200mM D-Ala, 10mM NADH, 90U/ml LDH, 200mM glucose, 30U/ml GDH, reaction at 30 ℃ for 15min, 30min, 1h, 2h, 3h, 4h, and termination of the reaction by addition of methanol after the reaction was completed. The reaction products were first separated using a common liquid phase column and the diastereomer selectivity was investigated, the analytical method being as described above. The retention times of (erythro) -and (threo) -1, 3-dihydroxy-1-p-methylsulfonylpropiophenone were 13.2 and 14.1min, respectively. According to the peak of the liquid phase, collecting a (threo) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylalkane sample, concentrating, further separating by a chiral liquid phase column, and inspecting the selectivity of the enantiomer. The conditions for the enantiomeric analysis were: chiral column IG column, column temperature 25 deg.C, 0.5ml/min, 224nm, chromatographic conditions: pure methanol (containing 0.1% diethylamine) for 15 min. The retention times of (1R,2R) -and (1S,2S) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylpropane were 5.6 and 9.4min, respectively. ee ═ [ (RR-SS)/(RR + SS) ] × 100%. Wherein: RR and SS represent (1R,2R) -and (1S,2S) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylpropane. The conversion rate and stereoselectivity of ATA117_ ACHH mutant protein catalytic synthesis of (1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylalkane are shown in the figures 2 and 3.

Although the present invention has been described with reference to the preferred embodiments, it should be understood that various changes and modifications can be made therein by those skilled in the art without departing from the spirit and scope of the invention as defined in the appended claims.

Sequence listing

<110> Shanghai university of transportation

<120> construction and application of enantioselectively inverted omega-transaminase mutant

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Gly Leu Asp Tyr Ile Thr Tyr Ser Asp Tyr Glu Leu Asp Pro Ala Asn

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Pro Leu Ala Gly Gly Ala Ala Trp Ile Glu Gly Ala Phe Val Pro Pro

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Ser Glu Ala Arg Ile Ser Ile Phe Asp Gln Gly Tyr Leu His Ser Asp

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Val Thr Tyr Thr Ala Phe His Val Trp Asn Gly Asn Ala Phe Arg Leu

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Asp Asp His Ile Glu Arg Leu Phe Ser Asn Ala Glu Ser Met Arg Ile

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Ile Pro Pro Leu Thr Gln Asp Glu Val Lys Glu Ile Ala Leu Glu Leu

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Val Ala Lys Thr Glu Leu Arg Glu Ala Cys Val Ser Val Ser Ile Thr

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Arg Gly Tyr Ser Ser Thr Pro Gly Glu Arg Asp Ile Thr Lys His Arg

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Pro Gln Val Tyr Met Tyr Ala Val Pro Tyr Gln Trp His Val Pro Phe

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Asp Arg Ile Arg Asp Gly Val His Ala Met Val Ala Gln Ser Val Arg

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Arg Thr Pro Arg Ser Ser Ile Asp Pro Gln Val Lys Asn Phe Gln Trp

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Gly Asp Leu Ile Arg Ala Val Gln Glu Thr His Asp Arg Gly Phe Glu

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Ala Pro Leu Leu Leu Asp Gly Asp Gly Leu Leu Ala Glu Gly Ser Gly

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Ser Ser Ser Leu Leu Thr Pro Val Gln Tyr

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atggcattta gcgcggatac cagcgaaatc gtctataccc acgataccgg tctggattac 60

atcacctaca gcgattacga actggatccg gcaaatccgt tagcaggcgg cgcagcttgg 120

attgaaggcg catttgttcc gccgtctgaa gcgcgtatta gcatcttcga tcagggctat 180

ctgcatagcg acgttaccta taccgcattc cacgtttgga acggtaacgc atttcgtctg 240

gacgatcaca tcgaacgtct gttcagcaac gccgaaagta tgcgtattat tccgccgctg 300

acccaagacg aagtcaaaga aatcgcgctg gaactggttg caaaaaccga actgcgcgaa 360

gcgtgtgtta gcgttagcat tacccgcggt tatagtagta ccccgggcga acgcgatatt 420

accaaacatc gtccgcaggt ctacatgtac gctgttccgt accagtggca tgttccgttt 480

gatcgtattc gcgacggcgt tcacgcaatg gttgcacaga gcgtgcgtcg taccccgcgt 540

agcagcatcg atccgcaggt caaaaacttc cagtggggcg atttaattcg cgcagttcag 600

gaaacccacg atcgcggttt tgaagcaccg ttactgttag acggcgacgg tttactggca 660

gaaggtagcg gccataacgt cgtcgtcatt aaagatgggg ttgttcgttc tccgggtcgt 720

gcagcattac cgggtattac ccgcaaaacc gttctggaaa ttgcggaatc cctgggtcat 780

gaagcgattc tggcggatat taccttagcg gaactgctgg acgcagacga agttttaggt 840

tgtaccaccg ctggcggcgt ttggccgttt gttagcgttg acggcaatcc gattagcgat 900

ggtgttccgg gtccggttac ccaaagcatt attcgtcgct actgggagct gaacgttgaa 960

agtagcagcc tgttaacccc ggttcagtat taa 993

Claims

1. An enantioselectively inverted omega-transaminase ATA117 mutant is characterized in that the protein amino acid sequence of the omega-transaminase ATA117 mutant is shown as SEQ ID No. 1.

2. The ω -transaminase ATA117 mutant according to claim 1, which is produced by iterative saturation mutation of the amino acid sequence of ω -transaminase ATA117, wherein the ω -transaminase ATA117 mutant has a mutation of at least one of valine at position N1, phenylalanine at position N2, isoleucine at position N3, phenylalanine at position N4.

3. The ω -transaminase ATA117 mutant according to claim 2, characterized by a mutation of valine at position N1 to alanine, a mutation of phenylalanine at position N2 to cysteine, a mutation of isoleucine at position N3 to histidine and a mutation of phenylalanine at position N4 to histidine.

4. A DNA nucleotide sequence comprising a nucleotide sequence encoding a ω -transaminase ATA117 mutant according to claim 2.

5. A DNA nucleotide sequence as claimed in claim 4, wherein the DNA nucleotide sequence is as shown in SEQ ID No. 2.

6. A plasmid carrying the DNA nucleotide sequence of claim 5.

7. A genetically engineered bacterium for expressing a omega-transaminase ATA117 mutant is characterized in that a protein amino acid sequence of the omega-transaminase ATA117 mutant is shown as SEQ ID No. 1.

8. The method for constructing genetically engineered bacteria expressing ω -transaminase ATA117 mutants according to claim 7, comprising the steps of: a1, using recombinant plasmid carrying gene coding ATA117 as template, A2, designing and synthesizing primer containing mutation, A3, reverse PCR amplifying whole plasmid, A4, PCR product transforming expression host.

9. The method for constructing genetically engineered bacteria expressing ω -transaminase ATA117 mutants according to claim 8, wherein the expression host is e.coli bl21(DE 3).

10. A method for producing (1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonylphenylpropane 2b based on the ω -transaminase ATA117 mutant as claimed in claim 1: the omega-transaminase ATA117 mutant pure protein is used as a catalyst to catalyze racemic 1, 3-dihydroxy-1-p-methylsulfonyl propiophenone 1 in a buffer system, and the (1R,2R) -1, 3-dihydroxy-2-amino-1-p-methylsulfonyl phenylpropane 2b with high stereoselectivity is obtained through kinetic resolution.