CN113683813A

CN113683813A - Application of dye decoloration peroxidase in degradation of polystyrene

Info

Publication number: CN113683813A
Application number: CN202110947698.8A
Authority: CN
Inventors: 吴敬; 夏伟; 杜妍怡; 姚从禹; 窦明德; 宿玲恰
Original assignee: Jiangnan University
Current assignee: Jiangnan University
Priority date: 2021-08-18
Filing date: 2021-08-18
Publication date: 2021-11-23
Anticipated expiration: 2041-08-18
Also published as: CN113683813B

Abstract

The invention discloses an application of dye decoloration peroxidase in polystyrene degradation, belonging to the technical field of enzyme engineering. The present invention provides decolorized dyes from Thermomonospora curvata and NostocaceaeUse of oxidases TcDyP and AnaPX for degrading Polystyrene (PS). The invention expresses dye decolorizing peroxidase TcDyP and AnaPX in colon bacillus, collects enzyme in fermentation liquor, adds the enzyme into glycine-HCl buffer solution containing PS film, uses H₂O₂Catalyzing, and reacting in constant-temperature water bath shaking tables with the temperature of 30 ℃ and the temperature of 25 ℃ and the speed of 200rpm for 48 hours to degrade the polystyrene plastic part, thereby having important application value for realizing the biodegradation of the plastic.

Description

Application of dye decoloration peroxidase in degradation of polystyrene

Technical Field

The invention relates to application of dye decoloration peroxidase in polystyrene degradation, belonging to the technical field of enzyme engineering.

Background

Plastics have been widely used in the fields of furniture, automotive parts, packaging, body implants, aviation and aerospace, and have become one of the most commonly used commodities in daily life. The global plastic consumption is over 3.2 hundred million tons per year, and the consumption is increased at a rate of 4-6% per year. Among them, Polystyrene (PS) is a plastic with light performance, heat resistance and low production cost. Up to now, styrofoam is sold free of charge in shops, stalls and even supermarkets. Is widely used in catering industry, electronic industry, medical field and the like, and accounts for about 6.6 percent of the total amount of plastics. Because it is difficult to degrade, this allows the PS plastic waste to continue to accumulate in the environment, which poses a serious ecological threat.

Currently, the degradation of Polystyrene (PS) still remains in the stage of using chemical physical degradation methods such as acid, alkali, alcohol, heat, etc. Among them, the chemical degradation method may require a large amount of chemicals, high temperature and high pressure equipment, and may generate a large amount of toxic and harmful substances, which may have a serious influence on the ecological environment. Thus, biodegradation is considered to be the most effective method for controlling plastic contamination.

Because the chemical degradation method used at present has the problems of high cost, serious three-waste pollution and the like, a biological method which is green, pollution-free and low in cost is increasingly used in a plurality of manufacturing fields, and the method gradually becomes a research hotspot in the field of plastic degradation. Currently, reported PS degrading bacteria mainly comprise cephalosporium, acinetobacter, tenebrio molitor intestinal flora and the like, but PS degrading enzymes are not reported yet. Therefore, there is a need to find PS degrading enzymes capable of degrading PS plastics.

Disclosure of Invention

The first object of the present invention is to provide a novel use of a dye decolorizing peroxidase which is (a) or (b) or (c):

(a) NCBI accession numbers of amino acid sequences such as ACY98529.1 or WP _010995754.1 dye decolorizing peroxidase;

(b) the amino acid sequence is shown as SEQ ID NO.1 or SEQ ID NO. 2;

(c) protein derived from (a) or (b) by substituting, deleting or adding one or more amino acids in the amino acid sequence defined by (a) or (b) and having dye decolorizing peroxidase activity.

In one embodiment, the dye decolorizing peroxidase is used in the degradation of polystyrene in the form of a free, whole cell, or physically/chemically immobilized enzyme.

In one embodiment, the degradation is a degradation reaction of dye decolorizing peroxidase with an amino acid sequence shown as SEQ ID No.1 or 2 in a reaction system containing Polystyrene (PS).

In one embodiment, the amount of the dye decolorizing peroxidase added in the reaction system is more than or equal to 1X 10³U/L。

In one embodiment, the dye decolorizing peroxidase represented by SEQ ID NO.1 is added in an amount of 2.6X 10 or more³U/L。

In one embodiment, the dye decolorizing peroxidase represented by SEQ ID NO.2 is added in an amount of 3.0X 10 or more⁴U/L。

In one embodiment, the temperature of the degradation reaction is 20-35 ℃, or 20-25 ℃, or 25-30 ℃, or 25-35 ℃, or 30-35 ℃.

In one embodiment, the reaction system has a pH of 3 to 4.

In one embodiment, the reaction system further comprises 50mM glycine-HCl buffer.

In one embodiment, the degradation temperature is 30 ℃ and 25 ℃ and the degradation time is 48 h.

In one embodiment, the degradation system is a glycine-HCl buffer at pH3 and pH3.5, 50 mM.

In one embodiment, the degradation reaction further comprises subjecting the polystyrene to an ultraviolet pretreatment prior to enzymatic hydrolysis; the ultraviolet pretreatment is to expose the polystyrene to ultraviolet rays for a certain time.

In one embodiment, the pre-treatment is performed by irradiating the polystyrene with ultraviolet light at 275-.

The invention also provides a gene for coding the dye decolorizing peroxidase.

In one embodiment, the nucleotide sequence encoding the dye decolorizing peroxidase represented by SEQ ID NO.1 is represented by SEQ ID NO. 3; the nucleotide sequence of the dye decolorizing peroxidase shown in the code SEQ ID NO.2 is shown in the SEQ ID NO. 4.

The invention also provides an expression vector carrying the dye decolorizing peroxidase gene.

In one embodiment, the expression vector is a PET vector.

In one embodiment, the expression vector is a PET-24a (+) plasmid.

The invention also provides a microbial cell containing the expression vector or expressing the dye decolorizing peroxidase.

In one embodiment, the host cell is E.coli.

In one embodiment, the host cell is Escherichia coli BL21

The invention also provides application of the gene or the expression vector or the cell in producing the dye decolorizing peroxidase.

In one embodiment, the production of the dye-decolorizing peroxidase is performed by culturing a microbial cell expressing the dye-decolorizing peroxidase in a culture medium and collecting the dye-decolorizing peroxidase.

Has the advantages that:

(1) the invention provides a new gene for coding heterologous expression dye decolorizing peroxidase, constructs recombinant escherichia coli for heterologous expression dye decolorizing peroxidase, and realizes heterologous expression of dye decolorizing peroxidase from Thermomonospora curvata and Nostocaceae.

(2) The invention provides a new application of dye decolorizing peroxidase in degrading polystyrene. Dye decolorizing peroxidase was added to Polystyrene (PS) -containing films (2 x 2 cm)²) The reaction system of (2) can be respectively reacted for 48 hours at 30 ℃, pH3 and 25 ℃, pH3.5, so that the Polystyrene (PS) can be partially degraded, and the surface of the film is obviously corroded. The method has mild reaction conditions and environmental protection, and has extremely high application prospect for realizing the biodegradation of the Polystyrene (PS) plastic.

Drawings

FIG. 1: SDS-PAGE analysis of dye decolorizing peroxidase; wherein (a) AnaPX; (b) TcDyP.

FIG. 2: polystyrene (PS) pre-oxidized by uv and cleaned and changed by ir.

FIG. 3: the dye decolorization peroxidase changes the microstructure and hydrophilic angle of the surface of a Polystyrene (PS) film. Wherein (a) is SEM; (b) a hydrophilic angle.

FIG. 4: dye decolorization peroxidase changes the surface of Polystyrene (PS) thin surface in infrared.

Detailed Description

The invention is further illustrated with reference to specific examples.

The Polystyrene (PS) films referred to in the examples below were obtained from Goodfellow GmbH, product No. ST311030, thickness month 0.030 mm; dye decolorization peroxidase tcdyp (thermomonospora curvata) and dye decolorization peroxidase anapx (cyanobacterium) are expressed by way of example.

The media involved in the following examples are as follows:

LB solid Medium (g/L): peptone 10, yeast powder 5, sodium chloride 10, agar 13, pH 7.0.

LB liquid Medium (g/L): peptone 10, yeast powder 5, sodium chloride 10, pH 7.0.

The detection methods referred to in the following examples are as follows:

the detection method of the microstructure on the surface of the Polystyrene (PS) plastic film comprises the following steps:

repeatedly washing the Polystyrene (PS) film treated by dye decolorizing peroxidase with SDS for 2 times, and then washing with deionized water for 1 time; putting the cleaned Polystyrene (PS) film into an oven for drying at 60 ℃; and (3) detecting the microstructure of the surface of the untreated Polystyrene (PS) film and the microstructure of the surface of the Polystyrene (PS) film after dye decoloration peroxidase treatment by using the untreated Polystyrene (PS) film as a reference through a scanning electron microscope.

The detection method of the surface of the Polystyrene (PS) film comprises the following steps:

repeatedly washing the Polystyrene (PS) film treated by dye decolorizing peroxidase with SDS for 2 times, and then washing with deionized water for 1 time; putting the cleaned Polystyrene (PS) film into an oven to be dried at 60 DEG C

The dried PS film is subjected to Scanning Electron Microscopy (SEM), hydrophilic angle and infrared spectroscopy (ATR-FT-IR), and the detection method refers to the article Interaction between Styrofoam and Microalgae spiral tissue in Brackish Water System published in 2021.

The method for measuring the enzyme activity of the dye decolorizing peroxidase comprises the following steps: enzyme activity assay is carried out at optimum pH and optimum temperature in 870. mu.L glycine-HCL buffer (100mM), 10. mu.L hydrogen peroxide (1mM), 20. mu.L ABTS (0.2mM), and 100. mu.L enzyme solution; wherein the optimal temperature of the dye decolorizing peroxidase TcDyP is 30 ℃, and the optimal pH is 3; the optimal temperature of dye decolorizing peroxidase AnaPX is 25 ℃, and the optimal pH is 3.5.

Definition of enzyme activity: the amount of enzyme required to oxidize 1. mu. mol of 2, 2' -azo-bis (3-ethylbenzothiazoline-6-sulfonic Acid (ABTS)) per minute in the reaction system.

Technical terms:

dye decolorizing peroxidase: the term "dye-decolorizing peroxidase" (dye-decolorizing peroxidase) is a protein containing heme that degrades various toxic dyes and can participate in the regulation of oxidative stress of bacteria and increase the resistance of bacteria to oxidative stress. Taking the embodiment of the invention as an example, the dye decolorizing peroxidase is derived from Thermomonospora curvata (Thermomonospora curvata) and has an amino acid sequence shown as NCBI accession number ACY98529.1 or shown as SEQ ID NO.1, or the dye decolorizing peroxidase is derived from Nostocaceae (Nostocaceae) and has an amino acid sequence shown as NCBI accession number WP _010995754.1 or shown as SEQ ID NO. 2.

Gene encoding dye decolorizing peroxidase: the term "gene encoding dye decolorizing peroxidase" directly specifies a polynucleotide encoding dye decolorizing peroxidase, which is typically bounded by an open reading frame beginning with a start codon (e.g., ATG, GTG, or TTG) and ending with a stop codon (e.g., TAA, TAG, or TGA).

Expressing: the term "expression" includes any step involved in the production of dye decolorization peroxidase, including but not limited to transcription, post-transcriptional modification, translation, post-translational modification, and secretion.

Expression vector: the term "expression vector" means a linear or circular DNA molecule comprising a gene encoding the dye decolorizing peroxidase of the present invention and operably linked to control sequences providing for its expression.

Host cell: the term "host cell" means any cell type that is susceptible to transformation, transfection, transduction, and the like with a nucleic acid construct or expression vector comprising a polynucleotide of the present invention. The term "host cell" encompasses any progeny of a parent cell that is not identical to the parent cell due to mutations that occur during replication.

Fermentation liquor: the term "fermentation broth" refers to a preparation produced by fermentation of cells, either unretained or recovered and/or purified. For example, a fermentation broth is produced when a microbial culture is grown to saturation by incubation under carbon-limited conditions that allow protein synthesis (e.g., expression of an enzyme by a host cell) and secretion of the protein into the cell culture medium. The fermentation broth may contain the contents of the fermented material obtained at the end of the fermentation. For example, the fermentation broth comprises the medium components after being utilized by the microorganism and cell debris present after the microbial cells have been removed by centrifugation.

Example 1: construction of recombinant plasmid carrying dye decolorization peroxidase gene TcDyP

Synthesizing a gene which has a nucleotide sequence shown as SEQ ID NO.3 and codes dye decolorizing peroxidase on a carrier pET-24a (+) by a chemical method, directly obtaining a recombinant plasmid, and transforming Escherichia coli (Escherichia coli) JM109 with the recombinant plasmid to obtain a transformation product; coating the transformation product on an LB solid culture medium (containing 40 mu g/mL kanamycin), and carrying out inverted culture in a constant-temperature incubator at 37 ℃ for 8-12 h to obtain a transformant; and (3) selecting a transformant, inoculating the transformant to an LB liquid culture medium, performing shake-flask culture for 8-12 h at 37 ℃ and 120-180 rpm, extracting plasmids, performing sequencing verification, and obtaining the recombinant plasmid pET-24a (+) -TcDyP after verification is correct.

Example 2: construction of recombinant plasmid carrying dye decolorizing peroxidase gene AnaPX

Synthesizing a gene which has a nucleotide sequence shown as SEQ ID NO.4 and codes dye decolorizing peroxidase on a carrier pET-24a (+) by a chemical method, directly obtaining a recombinant plasmid, and transforming Escherichia coli (Escherichia coli) JM109 with the recombinant plasmid to obtain a transformation product; coating the transformation product on an LB solid culture medium (containing 40 mu g/mL kanamycin), and carrying out inverted culture in a constant-temperature incubator at 37 ℃ for 8-12 h to obtain a transformant; and (3) selecting a transformant, inoculating the transformant to an LB liquid culture medium, performing shake-flask culture for 8-12 h at 37 ℃ and 120-180 rpm, extracting plasmids, performing sequencing verification, and obtaining the recombinant plasmid pET-24a (+) -AnaPX after verification is correct.

Example 3: construction of recombinant Escherichia coli expressing dye decolorizing peroxidase

Transforming the recombinant plasmid into Escherichia coli (Escherichia coli) BL21 to obtain a transformation product; coating the transformation product on an LB solid culture medium (containing 50 mu g/mL kanamycin), and carrying out inverted culture in a constant-temperature incubator at 37 ℃ for 8-12 h to obtain a transformant; and (3) selecting a transformant, inoculating the transformant to an LB liquid culture medium, performing shake flask culture for 8-12 h at 37 ℃ and 120-180 rpm, extracting plasmids, performing enzyme digestion verification and sequencing verification, and obtaining the recombinant escherichia coli BL21-pET-24a (+) -TcDyP after verification is correct.

Example 4: construction of recombinant Escherichia coli expressing dye decolorizing peroxidase

Transforming the recombinant plasmid into Escherichia coli (Escherichia coli) BL21 to obtain a transformation product; coating the transformation product on an LB solid culture medium (containing 50 mu g/mL kanamycin), and carrying out inverted culture in a constant-temperature incubator at 37 ℃ for 8-12 h to obtain a transformant; and (3) selecting a transformant, inoculating the transformant to an LB liquid culture medium, performing shake flask culture for 8-12 h at 37 ℃ and 120-180 rpm, extracting plasmids, performing enzyme digestion verification and sequencing verification, and obtaining the recombinant escherichia coli BL21-pET-24a (+) -AnaPX after verification is correct.

Example 5: recombinant expression of dye decolorizing peroxidase

Respectively culturing the recombinant Escherichia coli BL21-pET-24a (+) -TcDyP obtained in the construction of the example 3 and the recombinant Escherichia coli BL21-pET-24a (+) -AnaPX obtained in the construction of the example 4 in a shake flask containing an LB culture medium for 8-12 h, respectively transferring the bacterial liquid with the inoculation amount of 5% (v/v) to 100mL of LB liquid culture medium, and shake-culturing at 37 ℃ and 200rpm for 3h to OD₆₀₀After the concentration is 0.8, IPTG with a final concentration of 0.4mM and 5-aminolevulinic acid with a final concentration of 2mM are added, and recombinant escherichia coli BL21-pET-24a (+) -TcDyP is subjected to shaking culture at 30 ℃ and 200rpm for 20 hours to obtain TcDyP fermentation liquor (OD is about 7-10); BL21-pET-24a (+) -AnaPX is continuously subjected to shake cultivation at 25 ℃ and 200rpm for 20h to obtain AnaPX fermentation liquor (OD is about 7-10).

Respectively centrifuging fermentation liquor obtained by fermentation for 15min at 8000rpm to obtain precipitate; suspending the precipitate with a pH 7.0100 mM potassium phosphate buffer to obtain a resuspension; breaking the wall of the heavy suspension by using a high-pressure homogenizer under the pressure of 800Bar to obtain a wall-broken liquid; centrifuging the wall-broken solution at 8000rpm for 15min, and separating the wall-broken supernatant and the wall-broken precipitate.

The enzyme activity of the dye decolorizing peroxidase was measured separately (as shown in Table 1). SDS-PAGE analysis of the wall-broken supernatant showed that there were crude protein bands at the 41.9kDa and 54.3kDa bands, i.e., TcDyP and AnaPX, which are the dye decolorizing peroxidases (shown in FIG. 1).

TABLE 1 dye decolorizing peroxidase enzyme Activity

Example 6: ultraviolet pre-oxidation and cleaning of Polystyrene (PS) film

The method comprises the following specific steps:

polystyrene (PS) films were irradiated for 8 days at UVC (275-200 nm). The film was placed 20 cm from the lamp and turned over each day to allow each side of the samples to be uniformly illuminated. Cutting the UV-irradiated 8-day PS film into 2 × 2cm²And the size of the membrane is that 2 times of ultrasonic washing is carried out by 2% SDS, then deionized water is used for washing twice, then 75% ethanol is used for 2 times of ultrasonic washing and deionized water is used for washing twice, finally n-hexane is used for standing and washing for 2 hours, then deionized water is used for washing twice, and the residual n-hexane on the membrane is thoroughly removed and then dried to be used as an enzyme degradation substrate for later use. And performing infrared spectrum measurement on the dried PS film.

The infrared spectrum of the Polystyrene (PS) subjected to ultraviolet pre-oxidation and washing is detected, and the result shows that the Polystyrene (PS) film can be oxidized by the ultraviolet pre-oxidation, oxygen-containing functional groups such as C ═ O, -OH and the like are inserted into the Polystyrene (PS) film, so that an action target point is generated for the subsequent dye decolorizing peroxidase to attack the Polystyrene (PS) film, and meanwhile, some small-molecule oxidation products are generated, but the small-molecule oxidation products can be removed after washing, and the influence of the small-molecule oxidation products on the subsequent degradation of the Polystyrene (PS) film by the dye decolorizing peroxidase is avoided (the detection result is shown in fig. 2).

Example 7: degradation effect of dye decolorizing peroxidase on Polystyrene (PS) film

(1) Degradation of polystyrene by dye decoloration peroxidase TcDyP

A Polystyrene (PS) film (2X 2 cm) treated in example 6 was used²) After placing in a 50mL centrifuge tube, the reaction system was adjusted to pH3.5 with potassium phosphate buffer (pH 3.5 and pH 2,100mM) and added to a final concentration of 2.6X 10³U/L of the peroxidase solution TcDyP for decolorization of dye obtained in example 5, to which H was added₂O₂(2mM and 1 mM). Reacting for 48 hours at 30 ℃, pH3 and 200rpm, separating the membrane from the degradation liquid, cleaning and drying the surface of the membrane, and carrying out SEM (scanning Electron microscope), hydrophilic angle and infrared spectrum measurement on the dried PS film.

(2) Degradation of polystyrene by dye decoloration peroxidase AnaPX

A Polystyrene (PS) film (2X 2 cm) treated in example 6 was used²) After being placed in a 50mL centrifuge tube, the dye decolorizing peroxidase AnaPX was adjusted to pH3 with potassium phosphate buffer (pH 3.5 and pH 2,100mM) at 3.0X 10⁴U/L addition amount the dye decolorization peroxidase AnaPX enzyme solution obtained in example 5 was added, and H was added₂O₂(2mM and 1 mM). Reacting at 25 ℃, pH3.5 and 200rpm for 48 hours, separating the membrane from the degradation liquid, cleaning and drying the surface of the membrane, and carrying out SEM, hydrophilic angle and infrared spectrum measurement on the dried PS film.

The detection of the surface microstructure and the change of the hydrophilic angle of the Polystyrene (PS) treated with the dye decolorization peroxidase solution shows that the dye decolorization peroxidases TcDyP and AnaPX have an etching effect on the surface of the Polystyrene (PS) film and can improve the hydrophilicity of the surface of the Polystyrene (PS) film (see the detection result in fig. 3). The infrared spectrum of the Polystyrene (PS) treated with the dye decolorization peroxidase solution is detected, and the result shows that the dye decolorization peroxidase TcDyP and AnaPX can oxidize the Polystyrene (PS) film to further generate oxygen-containing functional groups such as C ═ O, -OH, C-O-C, and the like, so that the Polystyrene (PS) is depolymerized (see fig. 4).

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.

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gccttcaagt acaggcagaa gggcgtgagc ggcgacgtgt tcgccaactt cttcctgagc 300

aggcacggct acgagtacct ggagatcgag cccttccaga tccccggcga caagcccttc 360

aggatgggca tgaagaacga ggagatcagg agcagcctgg gcgaccccaa gatcgccacc 420

tgggagctgg gcttccagag cgagatccac gccctggtgc tgatcgccga cgacgacatc 480

gtggacctgc tgcagatcgt gaaccagatc acccagaagc tgaggcagat cgccgagatc 540

gtgcacaggg aggacggctt catcctgagg aaccaggccg gccagatcat cgagcacttc 600

ggcttcgtgg acggcgtgag ccagcccctg ttcatgaaga gggacgtggt gagggagagg 660

gtgaacaact gcgacttcga caagtgggac cccaaggccc ccctggacag catcctggtg 720

gaggacccca acggcaacac caaggacagc tacggcagct acctggtgta caggaagctg 780

gagcagaacg tgaaggcctt cagggaggac cagaggaagc tggcccagaa gctgaacatc 840

caggagaacc tggccggcgc cctgatcgtg ggcaggttcg ccgacggcac ccccgtgacc 900

ctgagcgaca tccccaccta cgccgtgacc cccaccaaca acttcaacta cgacggcgac 960

ctggccgcca ccaagtgccc cttccacagc cacaccagga agaccaaccc caggggcgac 1020

accgccaggc tgctgaccac cgacggccac ttcgacgagg ccttcaagga ggagaggggc 1080

cacaggatca ccaggagggc cgtgagctac ggcgagaaca accccagcaa ggagcccgtg 1140

agcggcagcg gcctgctgtt cctgtgcttc cagagcaaca tcgagaacca gttcaacttc 1200

atgcagagca ggtgggccaa cccccagaac ttcgtgcagg tgaacaccgg ccccgacccc 1260

ctgatcggcc agcccagcgg cacccagaag tggcccaaga agtggggcga gcccgagacc 1320

gaggagtaca acttccagct gtggatcaac atgaagggcg gcgagtactt cttcgccccc 1380

agcatcagct tcctgaagac cctggcccac caccaccacc accactga 1428

Claims

1. Use of a dye decolorizing peroxidase in the degradation of polystyrene or polystyrene-containing products, characterized in that said dye decolorizing peroxidase is (a) or (b) or (c):

(b) the amino acid sequence is shown as SEQ ID NO.1 or SEQ ID NO. 2;

2. The use according to claim 1, wherein the dye decolorizing peroxidase is used in the degradation of polystyrene in the form of a free, whole cell, or physically/chemically immobilized enzyme.

3. A method for degrading polystyrene is characterized in that dye decoloration peroxidase is reacted in a reaction system containing polystyrene; the addition amount of the dye decolorizing peroxidase in the reaction system is more than or equal to 1 multiplied by 10³U/L；

The dye decolorizing peroxidase is (a) or (b) or (c):

(b) the amino acid sequence is shown as SEQ ID NO.1 or SEQ ID NO. 2;

4. The method according to claim 3, wherein the temperature of the degradation reaction is 20 to 35 ℃.

5. The method according to claim 3 or 4, wherein the reaction system has a pH of 3 to 4.

6. The method according to any one of claims 3 to 5, wherein before the enzymatic hydrolysis, the polystyrene is further subjected to an ultraviolet pretreatment; the ultraviolet pretreatment is to expose the polystyrene to ultraviolet rays for a certain time.

7. A gene for coding dye decolorizing peroxidase, which is characterized in that the nucleotide sequence of the dye decolorizing peroxidase coded by SEQ ID NO.1 is shown as SEQ ID NO. 3; the nucleotide sequence of the dye decolorizing peroxidase shown in the code SEQ ID NO.2 is shown in the SEQ ID NO. 4.

8. An expression vector carrying the gene of claim 7.

9. A microbial cell comprising the expression vector of claim 8 or expressing the dye decolorizing peroxidase represented by SEQ ID No.1 or 2.

10. Use of the microbial cell of claim 9 for the production of a dye decolorizing peroxidase.