CN111662903B - Logarithmic phase specific promoter and application thereof - Google Patents

Abstract

The present invention provides a logarithmic phase-specific promoter (sequence shown in SEQ ID NO: 1‑10) and applications thereof. The logarithmic phase-specific induction promoter provided by the present invention is mainly focused on the logarithmic phase of bacterial growth to initiate recombinant expression, which is of great significance for the fermentation and production of lysine decarboxylase. The logarithmic phase of the protection of the present invention starts The seed can induce the expression of the lysine decarboxylase gene after the growth of the strain passes through the stagnation period, thus avoiding the pressure that the bacterium begins to express the gene during the stagnation period and brings the pressure on the bacterium to adapt to the fermentation environment, and can be used in the same fermentation Express more lysine decarboxylase over time. The invention provides powerful technical support for expressing lysine decarboxylase by microorganisms and using lysine decarboxylase as an in vitro enzyme to catalyze L-lysine to produce 1,5-pentanediamine.

Description

Log phase specific promoter and its application

technical field

The invention belongs to the field of biotechnology, and in particular relates to a logarithmic phase-specific promoter and its application.

Background technique

Lysine decarboxylase (LDC for short, classification number EC 4.1.1.18) widely exists in microorganisms, animals and higher plants, which can remove a carboxyl group from L-lysine to generate 1,5-pentanedi Amines (cadaverine) and CO ₂ . 1,5-Pentanediamine has a wide range of uses, for example, it can be polymerized with dibasic acids to synthesize new polyamides, which has high application value in industrial production. At present, the microbial production of pentamethylenediamine mainly adopts microbial fermentation production and microbial in vitro enzyme catalysis to produce pentamethylenediamine.

When catalyzing the production of 1,5-pentanediamine by microorganisms in vitro, the lysine decarboxylase gene expression cassette is often constructed based on the lactose operon, and the lactose operon with the lack of function of lacI is used, and the lac promoter can be constitutive Expression does not depend on the addition of inducers, and realizes the massive expression of heterologous genes accompanied by bacterial growth.

In addition to the lac promoter that depends on the lactose operon, it is also hoped to obtain some new promoters that can induce the transcription of downstream genes in the logarithmic phase of bacterial growth, and the induction of these promoters does not require the addition of inducers.

Contents of the invention

The purpose of the present invention is to provide a logarithmic phase-specific promoter and its application.

Another object of the present invention is to provide a method for whole-cell catalytic production of 1,5-pentanediamine.

In order to achieve the purpose of the present invention, the logarithmic phase-induced promoter provided by the inventors can use microorganisms to express a large amount of heterologous proteins in logarithmic phases, and provide technical support for using microorganisms to express proteins that are toxic to bacterial growth. Further, in order to realize whole-cell catalytic production of 1,5-pentanediamine, the gene encoding lysine decarboxylase was constructed behind a logarithmic phase-specific promoter, and transformed into a bacterial strain, and the resulting recombinant strain could be used Catalyzes the production of 1,5-pentanediamine in whole cells.

In the first aspect, the logarithmic phase-specific promoter provided by the present invention has a size of 37-42 bp, and at least includes the -10 region, the RNA polymerase ^бs specific recognition site and the -35 region;

Wherein, the position of the 3' end base of the promoter sequence corresponding to the transcription start site of the downstream gene is recorded as position -1, the -10 region is located at positions -12 to -7, and the base composition is 5' -TATACT-3';

The -13 base of the promoter sequence is C;

The RNA polymerase б ^s specific recognition site is located at positions -18 to -14, and the base composition is 5'-TTGTT-3';

The base composition of the -35 region is 5'-T(C or T)(C or T)(C or T)G(C or T or A)(T or C)-3', but 5'- Except for TCCCGCC-3', and the number of bases between the -35 region and the -10 region is 15-20bp.

Further, the sequence of the logarithmic phase-specific promoter is:

i) the nucleotide sequence shown in any one of SEQ ID NO: 1-10; or

ii) The nucleotide sequence shown in any one of SEQ ID NO: 1-10 is substituted, deleted and/or increased by one or more nucleotides and exhibits promoter activity in a logarithmic phase-specific manner sequence; or

iii) a nucleotide sequence that hybridizes to any of the sequences shown in SEQ ID NO: 1-10 and exhibits promoter activity in a logarithmic phase-specific manner under stringent conditions, and the stringent conditions are in the presence of 0.1% SDS Hybridize in 0.1×SSPE or 0.1×SSC solution containing 0.1% SDS at 65°C, and wash the membrane with this solution; or

iv) A nucleotide sequence that has more than 90% homology with the nucleotide sequence of i), ii) or iii) and exhibits promoter activity in a log-phase specific manner.

In a second aspect, the present invention provides biological materials containing the log-phase promoter, including but not limited to recombinant DNA, expression cassettes, transposons, plasmid vectors, phage vectors, viral vectors or engineering bacteria.

In a third aspect, the present invention provides any of the following applications of the logarithmic phase promoter:

1) Application as a logarithmic phase-specific promoter of prokaryotes;

2) Application in the construction of recombinant DNA, expression cassettes, transposons, plasmid vectors, phage vectors, viral vectors or engineering bacteria.

The prokaryote is a bacterium, such as bacteria in the genus Escherichia, Corynebacterium, Brevibacterium, Streptomyces, Hafnia; Preferably, the bacterium is selected from the group consisting of Escherichia coli (E.coli), Bacillus subtilis (B.subtilis), Streptomyces coelicolor (S.coelicolor), Hafnia alvei (H.alvei), Corynebacterium glutamicum Bacillus (C. glutamicum), or strains or genetically engineered bacteria after mutagenesis or random mutation.

Further, the present invention provides the log phase promoter as Escherichia coli (E.coli), Bacillus subtilis (B.subtilis), Streptomyces coelicolor (S.coelicolor), Hafnia alvei (H.alvei ) or Corynebacterium glutamicum (C.glutamicum) in the production of lysine decarboxylase in the fermentative production of lysine decarboxylase gene expression of the use of logarithmic phase-specific promoter regulation.

In the fourth aspect, the present invention provides a recombinant DNA, which is formed by operably linking the promoter with a downstream target gene.

In the present invention, the target gene is selected from nucleic acids encoding proteins, nucleic acids encoding ribozymes, and nucleic acids encoding antisense RNAs; preferably, the proteins are enzymes, hormones, antibodies or growth factors; more preferably, the The enzyme is selected from the group consisting of oxidoreductases, transferases, hydrolases, lyases, isomerases, ligases. Further, the lyase is a decarboxylase; the decarboxylase is an amino acid decarboxylase, such as lysine decarboxylase, tyrosine decarboxylase, arginine decarboxylase, ornithine decarboxylase or glutamic acid decarboxylase enzyme. Still further, the lysine decarboxylase is derived from Escherichia coli, Bacillus subtilis, Bacillus halodurans, Streptomyces coelicolor, hive hafni Hafnia alvei, Corynebacterium glutamicum or Klebsiella oxytoca. Furthermore, the lysine decarboxylase is encoded by cadA gene, ldcC gene, haldc gene, a fragment of cadA gene, a fragment of ldcC gene or a fragment of haldc gene. Even more preferred is lysine decarboxylase encoded by the cadA gene.

In the fifth aspect, the present invention provides an expression vector, which contains the recombinant DNA.

In some embodiments, the expression vector carries an expression cassette containing a lysine decarboxylase gene, and the expression cassette is driven by the log-phase specific promoter to express the lysine decarboxylase gene.

In some embodiments, the expression vector also contains a backbone plasmid capable of autonomous replication in the host cell; preferably, the backbone plasmid is selected from pUC18, pUC19, pBR322, pACYC, pET, pSC101 and their derivative plasmids.

In the sixth aspect, the present invention provides a transformant, which is a host bacterium carrying the expression vector.

The host bacterium is selected from bacterial species in Escherichia (Escherichia), Corynebacterium (Corynebacterium), Brevibacterium (Brevibacterium), Streptomyces (Streptomyces), Hafnia (Hafnia); preferably Preferably, the host bacterium is selected from the group consisting of Escherichia coli (E.coli), Bacillus subtilis (B.subtilis), Streptomyces coelicolor (S.coelicolor), Hafnia hive (H.alvei), rod-shaped glutamic acid Bacillus (C. glutamicum), or strains or genetically engineered bacteria after mutagenesis or random mutation; more preferably, the host bacteria is Escherichia coli (E.coli) or Hafnia alvei (H.alvei).

In the seventh aspect, the present invention provides the use of the transformant in the fermentative production of amino acids, polypeptides or proteins.

In the eighth aspect, the present invention provides an engineering bacterium producing lysine decarboxylase, the starting strain of the engineering bacterium is Escherichia coli (E.coli) or haffney bacteria (H.alvei), and carries A plasmid for an amino acid decarboxylase gene expression cassette, the expression of the lysine decarboxylase gene is driven by the log phase specific promoter.

Preferably, the lysine decarboxylase gene is an endogenous cadA gene (SEQ ID NO: 11) from Escherichia coli, which encodes an Escherichia coli inducible lysine decarboxylase CadA (SEQ ID NO: 12).

Preferably, the log-phase-specific promoter is the promoter shown in any one of SEQ ID NO: 1, 2, 3, 4, or 6.

Further, the engineered bacterium producing lysine decarboxylase provided by the present invention contains the logarithmic phase-specific promoter, or carries a plasmid containing a lysine decarboxylase gene expression cassette, and the lysine Acid decarboxylase gene expression cassette Expression of the lysine decarboxylase gene is driven by the log-phase specific promoter.

In the ninth aspect, the present invention provides a method for fermentatively producing lysine decarboxylase, comprising culturing the engineering bacteria producing lysine decarboxylase in a fermentation medium.

In the tenth aspect, the present invention provides a method for fermentatively producing 1,5-pentanediamine, comprising using the engineering bacteria obtained in the ninth aspect to ferment and produce lysine decarboxylase, and then using the obtained lysine decarboxylase to catalyze L - Conversion of lysine to 1,5-pentanediamine.

Preferably, the engineered bacteria are fermented to obtain an enzyme fermentation liquid, and the enzyme fermentation liquid is mixed with L-lysine, L-lysine salt solution or L-lysine fermentation liquid to catalyze the conversion of L-lysine into 1,5-Pentanediamine.

Preferably, the L-lysine fermentation liquid is selected from the L-lysine fermentation stock solution, the concentrate or dilution of the L-lysine fermentation stock solution, and the sterilizing solution after the L-lysine fermentation stock solution removes bacteria. L-lysine fermented liquid and concentrate or diluted liquid of sterilized L-lysine fermented liquid. The L-lysine fermentation broth can be prepared by existing techniques, for example, cultivating L-lysine-producing engineering bacteria in a fermentation medium to obtain an L-lysine fermentation broth, see CN104762336A, or the L-lysine fermentation broth. Lysine fermentation broth was obtained commercially.

Preferably, the method also includes adding a coenzyme to the L-lysine fermentation broth, and the coenzyme is selected from one or more of pyridoxal, pyridoxal phosphate, pyridoxine, and pyridoxamine , more preferably pyridoxal 5'-phosphate.

Preferably, when catalyzing the conversion of L-lysine to 1,5-pentanediamine, the temperature of the reaction system is 25°C-55°C, more preferably 28°C-40°C.

By virtue of the above technical solutions, the present invention has at least the following advantages and beneficial effects:

The logarithmic phase-specific induction promoter provided by the present invention mainly focuses on the logarithmic phase of bacterial growth to initiate recombinant expression, which is of great significance for the fermentation and production of lysine decarboxylase, compared with the constitutive Plac promoter , the logarithmic phase promoter protected by the present invention can induce the expression of lysine decarboxylase gene after the growth of the bacterial strain crosses the stagnation period, avoiding that the bacterium begins to express the gene in the stagnation period and brings the bacterium to adapt to the fermentation environment. The pressure can express more lysine decarboxylase in the same fermentation time. The invention provides powerful technical support for expressing lysine decarboxylase by microorganism and catalyzing L-lysine to produce 1,5-pentanediamine by using lysine decarboxylase.

detailed description

The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention. Unless otherwise specified, the examples are all in accordance with conventional experimental conditions, such as Sambrook et al. Molecular Cloning Experiment Manual (Sambrook J & Russell DW, Molecular Cloning: a Laboratory Manual, 2001), or in accordance with the conditions suggested by the manufacturer's instructions.

The specific steps of PCR amplification, plasmid extraction, enzyme digestion, enzyme digestion product ligation, transformation, and condition parameters mentioned in the following examples were all carried out according to the conditions suggested in the instructions of the purchased relevant enzymes and reagents. The DNA polymerase used for PCR amplification, the restriction endonuclease used for digestion, the ligase used for ligation of the digested product, and Escherichia coli E. coli BL21 competent cells were all purchased from Treasure Bioengineering (Dalian) Co., Ltd. Plasmid extraction kits, DNA gel recovery kits, and PCR purification kits were purchased from Corning Life Sciences (Wujiang) Co., Ltd. with the trademark Axygen, and primers were purchased from Thermo Fisher Scientific (China) Co., Ltd. with the trademark INVITROGEN.

The Hafnia alvei Am607 (Hafnia alvei Am607) mentioned in the following examples has been preserved in the China Center for Type Culture Collection, address: Wuhan, China, Wuhan University, zip code 430072, preservation number CCTCC NO: M2018737, preservation date 2018 November 1st.

The conversion rate of L-lysine-1,5-pentanediamine in the reaction system of the following examples is the percentage of the molar ratio of 1,5-pentanediamine to the initial L-lysine, wherein L-lysine and 1 , The detection of 5-pentanediamine was detected by nuclear magnetic resonance (BRUKERULTRASHIEDTM400PLUS, Beckman NMR).

The plasmid transformation method described in the following examples is as follows: the ligation product was added to 100 μl of E. coli E. coli BL21 (DE3) competent cells, and then heat-shocked at 42° C. for 90 seconds after 20 minutes of ice bathing. Add 1ml of LB after incubation on ice for 5min. Apply to the corresponding resistant plates.

The primers used in the following examples are as follows:

Example 1 Promoter sequence expressed in logarithmic phase

The present invention designs and synthesizes 10 promoter sequences, the length of the promoter is 37-42bp, respectively as follows (5'-3', the last base at the 3' end of the promoter sequence corresponds to the start of transcription of the downstream gene The position of the starting site is -1 position), in which the double underline marks the -10 region (-12～-7), and the base composition of the -10 region is 5'-TATACT-3'; the extended -10 region is ( -13 position) Conserved base C; -18~-14 base composition is 5'-TTGTT-3', this region is necessary for the specific recognition of RNA polymerase б ^s ; the single underline is marked as possible -35 Region, where the conserved region found in the present invention, regulates the start time of the promoter, and its base sequence can be 5'-T(C/T)(C/T)(C/T)G(C/T/A )(T/C)-3', but its sequence cannot be 5'-TCCCGCC-3', and the base number of the spacer between the -35 region and the -10 region of the promoter can be 15-20 bp.

GTATTG(SEQ ID NO:1)；1. CAG TCTTGTC AAATTTGTAAAAATTGTTC

GTATTG (SEQ ID NO: 1);

GTATTG(SEQ ID NO:2)；2. CCAT TCCTGAC AAATTTTTAATATTGTTC

GTATTG (SEQ ID NO: 2);

GTATTG(SEQ ID NO:3)；3. AA TCTCGCC AAATTTCAATTTGTTC

GTATTG (SEQ ID NO: 3);

GTATTG(SEQ ID NO:4)；4. TAAG TCTTGCC AAATCCTTATTGTTC

GTATTG (SEQ ID NO: 4);

GTATTG(SEQ ID NO:5)；5. TCCTGAC AAATTTACAATATTGTTC

GTATTG (SEQ ID NO: 5);

GTATTG(SEQ ID NO:6)；6. TCCTGCC AAATCCGTAAATTTTGTTC

GTATTG (SEQ ID NO: 6);

GTATTG(SEQ ID NO:7)；7. CAC TTTCGTC AAATTTGTAATTATTGTTC

GTATTG (SEQ ID NO: 7);

GTATTG(SEQ ID NO:8)；8. TCTTGTT AAATTTGTAATTATTGTTC

GTATTG (SEQ ID NO: 8);

GTATTG(SEQ ID NO:9)；9. TTTTGCT AAATTTGTAAATTATTGTTC

GTATTG (SEQ ID NO: 9);

GTATTG(SEQ ID NO:10)。10. CGGAG TCTTGAC AATTGTAAATTATTGTTC

GTATTG (SEQ ID NO: 10).

Example 2 Construction of red fluorescent protein expression plasmids containing different promoters

The coding genes of mCherry red fluorescent protein are all synthesized by the method of gene assembly. The amino acid sequence of red fluorescent protein mCherry is SEQ ID NO:14, and its nucleotide sequence is SEQ ID NO:15 after codon optimization. Codon optimization and its DNA assembly operations refer to Hoover DM & Lubkowski J, Nucleic Acids Research 30:10, 2002.

The codon-optimized mCherry gene was PCR amplified using primers mCherry-F (SEQ ID NO: 16) and mCherry-R (SEQ ID NO: 17), and a ribose binding site was introduced upstream of the gene start codon (RBS, SEQID NO: 18); after cutting the gel to recover the DNA fragment of the target size, carry out the A reaction, and then connect it with the pMD18-T (purchased from Treasure Bioengineering Dalian Co., Ltd.) vector, and transform the ligated product into E.coli BL21 competent cells were screened on a plate containing ampicillin resistance at a final concentration of 100 μg/ml, cultured overnight at 37°C (16h, the same below), and several clones were picked and shaken and sent for sequencing. Select the clone with the correct sequencing sequence and the reverse connection of the mCherry gene into the lactose promoter plac to extract the plasmid to obtain the plasmid T-mCherry containing the mCherry gene. During the sequencing process, it was found that the clones whose mCherry gene sequencing was verified to be correct and positively inserted into plac could appear red on the plate. Thus, the cloned mCherry gene can be successfully expressed in Escherichia coli and the translation products are all functional proteins. However, the clone after the mCherry gene was reversely inserted into plac, although the sequencing verification was correct, the clone did not appear red, indicating that the plasmid T-mCherry can be used for subsequent verification of the start time of the promoter.

Using the plasmid T-mCherry as a template, first use primer T-mC-F1 (SEQ ID NO: 19) and primer T-mC-R1 (SEQ ID NO: 20) to carry out plasmid amplification, and use the PCR purification kit to purify the PCR product After purification, the template was digested with restriction endonuclease DpnI, transformed into E.coli BL21 cells, screened on a resistance plate containing ampicillin at a final concentration of 100 μg/ml, and cultured overnight at 37°C. Three single clones were randomly selected for sequencing identification. Select the correct clone to extract the plasmid, use the plasmid as a template, and use primer T-mC-F2 (SEQ ID NO:21) and primer T-mC-R2 (SEQ ID NO:22) to carry out PCR amplification of the plasmid, After the PCR product was purified using the PCR purification kit, the plasmid template was digested with the restriction endonuclease DpnI, and then transformed into E. coli E. Screened on penicillin resistance plates and cultured overnight at 37°C. Three single clones were randomly selected for sequencing identification. The clones with correct sequencing were selected to extract the plasmids to obtain the plasmid T-Psyn-mCherry with the RBS sequence and multiple cloning sites inserted before the mCherry gene.

Double-stranded DNA sequences containing the 10 promoters listed in Example 1 were respectively synthesized using gene sequence synthesis methods commonly used in the art, and enzyme cleavage sites of KpnI and ClaI were respectively connected to the 5' and 3' ends of the sequence, and KpnI was used to After double digestion with KpnI and ClaI, they were ligated into the plasmid T-Psyn-mCherry which was digested with KpnI and ClaI, respectively. Plasmids containing different promoters were obtained. These 10 plasmids were respectively transformed into E. coli E. coli BL21 competent cells to obtain strains containing these 10 plasmids, which were named BL21-1-BL21-10 respectively.

Example 3 Using red fluorescent protein to verify the start time of the promoter

Pick 4 clones from the 10 plasmid-transformed plates and place them in 600 μl LB liquid medium containing ampicillin at a final concentration of 100 μg/ml in a 96-well deep-well plate, and store them at 37°C, After culturing overnight in a shaker at 250 rpm, transfer 20 μl of the bacterial solution to fresh 600 μl LB liquid medium containing ampicillin at a final concentration of 100 μg/ml, and culture in a shaker at 37°C and 250 rpm for 2 hrs, and then Transfer 8 μl of the bacteria solution to the fresh 200 μl LB liquid medium containing ampicillin at a final concentration of 100 μg/ml in the microplate plate (set up four replicates), and place it in a microplate reader. The bacteria solution of the bacterial strain that does not contain red fluorescent protein is set as a negative control, and the negative control bacteria grow normally, but no fluorescence is detected. Set the temperature to 37°C and set the workflow: first, shake the bacteria for 15s, measure the OD _600nm value of the bacteria, then measure the fluorescence value of the bacteria under the conditions of 575nm excitation light and 610nm emission light, and finally shake the bacteria for 600s, the The process cycle is measured every 15 minutes, and the total time is 24hrs. After the measurement, the growth curve of the bacteria and the fluorescence curve of the unit OD (the ratio of the fluorescence value of the bacteria at a certain time point to the OD measured at this time point) curves were drawn respectively. According to the corresponding relationship between the two curves, it is obtained which stage of the growth of the bacteria is the time point when the fluorescence of the bacteria is significantly enhanced. The promoter contained in the clone whose fluorescence value began to increase significantly after the logarithmic phase of cell growth was determined to be a logarithmic phase-specifically induced promoter.

As can be seen from Table 1, when the recombinant bacterial strain containing the promoter of the present invention is cultivated for 360-480 minutes, the OD of the thalline grows rapidly, from about 0.2 to about 0.4, and the fluorescence value/OD of the thalline also increases. significantly increased, from about 80 units to about 120 units; as the culture time of the bacteria continued to prolong, the OD and fluorescence value/OD of the bacteria did not increase significantly, indicating that these promoters are all on the right side of the growth of the bacteria. Several phases begin to initiate transcription of downstream genes.

Table 1 OD value and fluorescence value/OD of each strain detected by microplate reader at different culture time

Example 4 The logarithmic phase promoter is used to express lysine decarboxylase for in vitro enzymatic production of 1,5-pentanediamine

1. Construction of plasmids and recombinant strains containing logarithmic promoters for inducible expression of lysine decarboxylase

Using the genome of Escherichia coli MG1655 K12 as a template, the primers cadA-F (SEQ ID NO:23) and cadA-R (SEQ ID NO:24) were used for the first round of amplification, and the target fragment was gel-cut and recovered as a template , and then use primers cadA-F2 (SEQ ID NO: 25) and cadA-R (SEQ ID NO: 24) for the second round of amplification, after gel cutting and recovery, the 5' end with HindIII restriction site and ribosome The cadA gene (SEQ ID NO: 13) of the binding site; the plasmid containing 10 kinds of promoters obtained in the extraction example 2 was digested with HindIII and XbaI, and the cadA gene (SEQ ID NO: 13) Connect to get T-1-cadA, T-2-cadA, T-3-cadA, T-4-cadA, T-5-cadA, T-6-cadA, T-7-cadA, T- 8-cadA, T-9-cadA, T-10-cadA these 10 cadA expression plasmids, these 10 plasmids were respectively transformed into hive Hafney strain Am607, and 10 recombinant strains were obtained, HA-1-cadA, HA-2-cadA, HA-3-cadA, HA-4-cadA, HA-5-cadA, HA-5-cadA, HA-6-cadA, HA-7-cadA, HA-8-cadA, HA- 9-cadA, HA-10-cadA.

It is known that in the absence of LacI repressor protein expression, the plac promoter can be a constitutive promoter, and its downstream genes are expressed along with the growth of the bacterium. In order to compare plac and the logarithmic phase-induced promoter of the present invention in Pros and cons in expressing heterologous proteins. At the same time, using the pUC18 plasmid as a template, primers plac-F (SEQ ID NO: 26) and plac-R (SEQ ID NO: 27) were used to amplify the plac constitutive promoter (SEQ ID NO: 28), KpnI, ClaI After double digestion, it was ligated with the same double digestion plasmid T-1-cadA to obtain plasmid T-plac-cadA. It was also transformed into Haffney strain Am607 to obtain HA-plac-cadA recombinant strain.

2. Fermentation and culture of recombinant strains

Three single clones of the above-mentioned recombinant strains were respectively picked into 5 mL of LB liquid medium, and the LB liquid medium contained tryptone (tryptone) 1% (w/v), yeast extract (yeast extract) 0.5% (w /v), NaCl 1% (w/v), and ampicillin 100 μg/mL, cultured at 37° C. for 24 hours to obtain enzyme fermentation liquid, and measure the OD _560nm of each strain’s enzyme fermentation liquid.

3. Catalyze the conversion of L-lysine into 1,5-pentanediamine

Get 500 μl of enzyme fermentation broth of each recombinant strain obtained in the above steps, add 500 μl of L-lysine hydrochloride aqueous solution (L-lysine concentration is 400g/L), and the final concentration is 0.05mM 5'-pyridine phosphate Pyridoxal, pH natural, reaction temperature 37°C, rotation speed 250rpm, react for 2h, after the reaction, centrifuge the reaction solution (12000rpm, 3 min), take 500μl of the reaction solution for detection and calculate the L-lysine of each reaction system - 1,5-pentanediamine conversion (Table 2).

Table 2 Detection of the levels of remaining L-lysine and converted 1,5-pentanediamine in each reaction system

As can be seen from Table 2, by comparing 10 logarithmic phase promoters in the present invention with constitutive plac promoters to induce expression of lysine decarboxylase, the results show that the present invention screened logarithmic phase promoters 1-10 Both can catalyze the conversion of L-lysine into 1,5-pentanediamine, and the conversion rate of lysine decarboxylase induced by logarithmic promoters 1, 2, 3, 4, and 6 is higher than that of constitutive plac Sons are more dominant, with conversion rates of 78%, 75%, 71%, 68% and 62%, respectively.

Example 5 The logarithmic phase promoter is used to express lysine decarboxylase for in vitro enzymatic production of 1,5-pentanediamine

The method of constructing a plasmid containing a logarithmic phase promoter to induce expression of lysine decarboxylase and the recombinant strain, and the method of fermenting and cultivating the recombinant strain are the same as in Example 4, the difference is that the conversion of L-lysine is catalyzed to generate 1,5- Pentylenediamine: The obtained recombinant strains HA-1-cadA, HA-2-cadA, HA-3-cadA, HA-4-cadA, HA-6-cadA and HA-plac-cadA were mixed with L -Lysine fermentation broth is mixed and converted into 1,5-pentanediamine, which specifically includes the following steps:

Take 600 μl of lysine fermentation stock solution (purchased from Kaiser (Jinxiang) Biomaterials Co., Ltd.), with a lysine content of 11%. Add 100 μl of the above-mentioned enzyme fermentation broth and 5`-pyridoxal phosphate with a final concentration of 0.05 mM to this lysine fermentation stock solution, the pH is natural, the reaction temperature is 38 ° C, the rotation speed is 250 rpm, and the reaction is 2 hours. After the reaction, the reaction solution is After centrifugation (12000 rpm, 3 min), 500 μl of the reaction solution was taken for detection and the conversion rate of L-lysine-1,5-pentanediamine in each reaction system was calculated (Table 3).

Table 3 Detection of the levels of remaining L-lysine and converted 1,5-pentanediamine in each reaction system

As can be seen from Table 3, the lysine decarboxylase induced and expressed by the logarithmic phase promoters 1, 2, 3, 4, and 6 screened by the present invention also obtained higher The L-lysine-1,5-pentanediamine conversion rate, the logarithmic phase promoter screened by the present invention promotes and simplifies the process of catalytic conversion of L-lysine to 1,5-pentanediamine.

Although the present invention has been described in detail with general descriptions and specific implementations above, it is obvious to those skilled in the art that some modifications or improvements can be made on the basis of the present invention. Therefore, the modifications or improvements made on the basis of not departing from the spirit of the present invention all belong to the protection scope of the present invention.

sequence listing

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agcttctttg aatatgcgct gggtgctgct gaagatattg ctaataagat caagcagacc 360

actgacgaat atatcaacac tattctgcct ccgctgacta aagcactgtt taaatatgtt 420

cgtgaaggta aatatacttt ctgtactcct ggtcacatgg gcggtactgc attccagaaa 480

agcccggtag gtagcctgtt ctatgatttc tttggtccga ataccatgaa atctgatatt 540

tccatttcag tatctgaact gggttctctg ctggatcaca gtggtccaca caaagaagca 600

gaacagtata tcgctcgcgt ctttaacgca gaccgcagct acatggtgac caacggtact 660

tccactgcga acaaaattgt tggtatgtac tctgctccag caggcagcac cattctgatt 720

gaccgtaact gccacaaatc gctgacccac ctgatgatga tgagcgatgt tacgccaatc 780

tatttccgcc cgacccgtaa cgcttacggt attcttggtg gtatcccaca gagtgaattc 840

cagcacgcta ccattgctaa gcgcgtgaaa gaaacaccaa acgcaacctg gccggtacat 900

gctgtaatta ccaactctac ctatgatggt ctgctgtaca acaccgactt catcaagaaa 960

acactggatg tgaaatccat ccactttgac tccgcgtggg tgccttacac caacttctca 1020

ccgattacg aaggtaaatg cggtatgagc ggtggccgtg tagaagggaa agtgattac 1080

gaaacccagt ccactcacaa actgctggcg gcgttctctc aggcttccat gatccacgtt 1140

aaaggtgacg taaacgaaga aacctttaac gaagcctaca tgatgcacac caccacttct 1200

ccgcactacg gtatcgtggc gtccactgaa accgctgcgg cgatgatgaa aggcaatgca 1260

ggtaagcgtc tgatcaacgg ttctattgaa cgtgcgatca aattccgtaa agagatcaaa 1320

cgtctgagaa cggaatctga tggctggttc tttgatgtat ggcagccgga tcatatcgat 1380

acgactgaat gctggccgct gcgttctgac agcacctggc acggcttcaa aaacatcgat 1440

aacgagcaca tgtatcttga cccgatcaaa gtcaccctgc tgactccggg gatggaaaaa 1500

gacggcacca tgagcgactt tggtattccg gccagcatcg tggcgaaata cctcgacgaa 1560

catggcatcg ttgttgagaa aaccggtccg tataacctgc tgttcctgtt cagcatcggt 1620

atcgataaga ccaaagcact gagcctgctg cgtgctctga ctgactttaa acgtgcgttc 1680

gacctgaacc tgcgtgtgaa aaacatgctg ccgtctctgt atcgtgaaga tcctgaattc 1740

tatgaaaaca tgcgtattca ggaactggct cagaatatcc acaaactgat tgttcaccac 1800

aatctgccgg atctgatgta tcgcgcattt gaagtgctgc cgacgatggt aatgactccg 1860

tatgctgcat tccagaaaga gctgcacggt atgaccgaag aagtttacct cgacgaaatg 1920

gtaggtcgta ttaacgccaa tatgatccctt ccgtacccgc cgggagttcc tctggtaatg 1980

ccgggtgaaa tgatcaccga agaaagccgt ccggttctgg agttcctgca gatgctgtgt 2040

gaaatcggcg ctcactatcc gggctttgaa accgatattc acggtgcata ccgtcaggct 2100

gatggccgct ataccgttaa ggtattgaaa gaagaaagca aaaaataa 2148

<210> 12

<211> 715

<212> PRT

<213> Escherichia coli

<400> 12

Met Asn Val Ile Ala Ile Leu Asn His Met Gly Val Tyr Phe Lys Glu

1 5 10 15

Glu Pro Ile Arg Glu Leu His Arg Ala Leu Glu Arg Leu Asn Phe Gln

20 25 30

Ile Val Tyr Pro Asn Asp Arg Asp Asp Leu Leu Lys Leu Ile Glu Asn

35 40 45

Asn Ala Arg Leu Cys Gly Val Ile Phe Asp Trp Asp Lys Tyr Asn Leu

50 55 60

Glu Leu Cys Glu Glu Ile Ser Lys Met Asn Glu Asn Leu Pro Leu Tyr

65 70 75 80

Ala Phe Ala Asn Thr Tyr Ser Thr Leu Asp Val Ser Leu Asn Asp Leu

85 90 95

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

100 105 110

Ile Ala Asn Lys Ile Lys Gln Thr Thr Asp Glu Tyr Ile Asn Thr Ile

115 120 125

Leu Pro Pro Leu Thr Lys Ala Leu Phe Lys Tyr Val Arg Glu Gly Lys

130 135 140

Tyr Thr Phe Cys Thr Pro Gly His Met Gly Gly Thr Ala Phe Gln Lys

145 150 155 160

Ser Pro Val Gly Ser Leu Phe Tyr Asp Phe Phe Gly Pro Asn Thr Met

165 170 175

Lys Ser Asp Ile Ser Ile Ser Val Ser Glu Leu Gly Ser Leu Leu Asp

180 185 190

His Ser Gly Pro His Lys Glu Ala Glu Gln Tyr Ile Ala Arg Val Phe

195 200 205

Asn Ala Asp Arg Ser Tyr Met Val Thr Asn Gly Thr Ser Thr Ala Asn

210 215 220

Lys Ile Val Gly Met Tyr Ser Ala Pro Ala Gly Ser Thr Ile Leu Ile

225 230 235 240

Asp Arg Asn Cys His Lys Ser Leu Thr His Leu Met Met Met Ser Asp

245 250 255

Val Thr Pro Ile Tyr Phe Arg Pro Thr Arg Asn Ala Tyr Gly Ile Leu

260 265 270

Gly Gly Ile Pro Gln Ser Glu Phe Gln His Ala Thr Ile Ala Lys Arg

275 280 285

Val Lys Glu Thr Pro Asn Ala Thr Trp Pro Val His Ala Val Ile Thr

290 295 300

Asn Ser Thr Tyr Asp Gly Leu Leu Tyr Asn Thr Asp Phe Ile Lys Lys

305 310 315 320

Thr Leu Asp Val Lys Ser Ile His Phe Asp Ser Ala Trp Val Pro Tyr

325 330 335

Thr Asn Phe Ser Pro Ile Tyr Glu Gly Lys Cys Gly Met Ser Gly Gly

340 345 350

Arg Val Glu Gly Lys Val Ile Tyr Glu Thr Gln Ser Thr His Lys Leu

355 360 365

Leu Ala Ala Phe Ser Gln Ala Ser Met Ile His Val Lys Gly Asp Val

370 375 380

Asn Glu Glu Thr Phe Asn Glu Ala Tyr Met Met His Thr Thr Thr Ser

385 390 395 400

Pro His Tyr Gly Ile Val Ala Ser Thr Glu Thr Ala Ala Ala Met Met

405 410 415

Lys Gly Asn Ala Gly Lys Arg Leu Ile Asn Gly Ser Ile Glu Arg Ala

420 425 430

Ile Lys Phe Arg Lys Glu Ile Lys Arg Leu Arg Thr Glu Ser Asp Gly

435 440 445

Trp Phe Phe Asp Val Trp Gln Pro Asp His Ile Asp Thr Thr Glu Cys

450 455 460

Trp Pro Leu Arg Ser Asp Ser Thr Trp His Gly Phe Lys Asn Ile Asp

465 470 475 480

Asn Glu His Met Tyr Leu Asp Pro Ile Lys Val Thr Leu Leu Thr Pro

485 490 495

Gly Met Glu Lys Asp Gly Thr Met Ser Asp Phe Gly Ile Pro Ala Ser

500 505 510

Ile Val Ala Lys Tyr Leu Asp Glu His Gly Ile Val Val Glu Lys Thr

515 520 525

Gly Pro Tyr Asn Leu Leu Phe Leu Phe Ser Ile Gly Ile Asp Lys Thr

530 535 540

Lys Ala Leu Ser Leu Leu Arg Ala Leu Thr Asp Phe Lys Arg Ala Phe

545 550 555 560

Asp Leu Asn Leu Arg Val Lys Asn Met Leu Pro Ser Leu Tyr Arg Glu

565 570 575

Asp Pro Glu Phe Tyr Glu Asn Met Arg Ile Gln Glu Leu Ala Gln Asn

580 585 590

Ile His Lys Leu Ile Val His His Asn Leu Pro Asp Leu Met Tyr Arg

595 600 605

Ala Phe Glu Val Leu Pro Thr Met Val Met Thr Pro Tyr Ala Ala Phe

610 615 620

Gln Lys Glu Leu His Gly Met Thr Glu Glu Val Tyr Leu Asp Glu Met

625 630 635 640

Val Gly Arg Ile Asn Ala Asn Met Ile Leu Pro Tyr Pro Pro Gly Val

645 650 655

Pro Leu Val Met Pro Gly Glu Met Ile Thr Glu Glu Ser Arg Pro Val

660 665 670

Leu Glu Phe Leu Gln Met Leu Cys Glu Ile Gly Ala His Tyr Pro Gly

675 680 685

Phe Glu Thr Asp Ile His Gly Ala Tyr Arg Gln Ala Asp Gly Arg Tyr

690 695 700

Thr Val Lys Val Leu Lys Glu Glu Ser Lys Lys

705 710 715

<210> 13

<211> 1642

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 13

aagcgaacga acaacaagaa ggaaaacaag aacgagcaaa gaacacaggg ggaaaagaag 60

aacccaccgg aaccacgcgc gcgaacgcga accgagac ccgaacgacc ggacgacaaa 120

aacgacgaaa acaagcgcgc ggcggcgaga cgggaaaaaa accgagcggc gaagaaaagc 180

aaaagaacga gaaccgccgg acgcgcgcaa acgaccacccc gagaagccga agaccgcgac 240

agaagccgaa agcgcggggc gcgaagaagc aaaagacaag cagaccacga cgaaaacaac 300

acacgccccg cgacaaagca cgaaaagcgg aaggaaaaac cgacccggca cagggcggac 360

gcaccagaaa agcccggagg agccgcagac ggccgaaacc agaaacgaac cacagacgaa 420

cgggccgcgg acacagggcc acacaaagaa gcagaacaga acgccgcgca acgcagaccg 480

cagcacagggg accaacggac ccacgcgaac aaaagggaga ccgcccagca ggcagcacca 540

cgagaccgaa cgccacaaac gcgacccacc gagagagagc gagacgccaa caccgcccga 600

cccgaacgca cggacgggga cccacagagg aaccagcacg caccagcaag cgcggaaaga 660

aacaccaaac gcaaccggcc ggacagcgaa accaaccacc agaggcgcga caacaccgac 720

cacaagaaaa cacggaggaa accacacga cccgcggggg ccacaccaac ccaccgaacg 780

aaggaaagcg gagagcgggg ccggagaagg gaaaggaacg aaacccagcc accacaaacg 840

cggcggcgcc caggcccaga ccacgaaagg gacgaaacga agaaaccaac gaagccacag 900

agcacaccac caccccgcac acggacgggc gccacgaaac cgcgcggcga gagaaaggca 960

agcaggaagc gcgacaacgg cagaacggcg acaaaccgaa agagacaaac gcgagaacgg 1020

aacgaggcgg cgagaggcag ccggacaacg aacgacgaag cggccgcgcg cgacagcacc 1080

ggcacggcca aaaacacgaa acgagcacag acgacccgac aaagcacccg cgacccgggg 1140

aggaaaaaga cggcaccaga gcgacggacc ggccagcacg ggcgaaaacc cgacgaacag 1200

gcacgggaga aaaccggccg aaaccgcgcc gcagcacgga cgaaagacca aagcacgagc 1260

cgcgcggccg acgacaaacg gcgcgaccga accgcgggaa aaacagcgcc gccgacggaa 1320

gaccgaacag aaaacagcga caggaacggc cagaaaccac aaacgagcac cacaacgccg 1380

gacgagacgc gcagaaggcg ccgacgagga agacccgagc gcaccagaaa gagcgcacgg 1440

agaccgaaga agaccgacg aaaggaggcg aaacgccaaa gaccccgacc cgccgggagc 1500

ccggaagccg gggaaagaca ccgaagaaag ccgccggcgg agccgcagag cgggaaacgg 1560

cgccacaccg ggcgaaaccg aacacgggca accgcaggcg aggccgcaac cgaaggagaa 1620

agaagaaagc aaaaaaaaca ga 1642

<210> 14

<211> 236

<212> PRT

<213> mushroom coral (mushroom coral)

<400> 14

Met Val Ser Lys Gly Glu Glu Asp Asn Met Ala Ile Ile Lys Glu Phe

1 5 10 15

Met Arg Phe Lys Val His Met Glu Gly Ser Val Asn Gly His Glu Phe

20 25 30

Glu Ile Glu Gly Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr

35 40 45

Ala Lys Leu Lys Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp

50 55 60

Ile Leu Ser Pro Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His

65 70 75 80

Pro Ala Asp Ile Pro Asp Tyr Leu Lys Leu Ser Phe Pro Glu Gly Phe

85 90 95

Lys Trp Glu Arg Val Met Asn Phe Glu Asp Gly Gly Val Val Thr Val

100 105 110

Thr Gln Asp Ser Ser Leu Gln Asp Gly Glu Phe Ile Tyr Lys Val Lys

115 120 125

Leu Arg Gly Thr Asn Phe Pro Ser Asp Gly Pro Val Met Gln Lys Lys

130 135 140

Thr Met Gly Trp Glu Ala Ser Ser Glu Arg Met Tyr Pro Glu Asp Gly

145 150 155 160

Ala Leu Lys Gly Glu Ile Lys Gln Arg Leu Lys Leu Lys Asp Gly Gly

165 170 175

His Tyr Asp Ala Glu Val Lys Thr Thr Tyr Lys Ala Lys Lys Pro Val

180 185 190

Gln Leu Pro Gly Ala Tyr Asn Val Asn Ile Lys Leu Asp Ile Thr Ser

195 200 205

His Asn Glu Asp Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ala Glu Gly

210 215 220

Arg His Ser Thr Gly Gly Met Asp Glu Leu Tyr Lys

225 230 235

<210> 15

<211> 711

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 15

atggtgtcta aaggcgagga agataatatg gcgattatca aagaatttat gcgttttaaa 60

gtgcatatgg aaggcagcgt gaatgggcat gagtttgaaa ttgaaggcga aggagaaggc 120

cgtccgtatg aaggcaccca gaccgctaaa ctgaaagtga ccaaaggcgg accactgccg 180

tttgcgtggg attctgag cccgcagttt atgtatggca gcaaagcgta tgtgaaacat 240

ccggcggata ttccggatta tctgaaactg agctttccgg agggcttcaa atgggaacgt 300

gtgatgaatt ttgaagatgg cggcgtggtg accgtgaccc aggatagcag cctgcaagac 360

ggcgaattca tttacaaggt gaagctgcgt ggcaccaact ttcccagcga tggcccggtg 420

atgcagaaaa agaccatggg ctgggaggcg agcagcgaac gtatgtaccc ggaggatggc 480

gcgctgaagg gcgaaattaa gcagcgtctg aagttaaaag atggtgggca ctatgatgcg 540

gaagtgaaaa ccacctataa agcgaaaaaa ccggtgcagt taccaggcgc ttataatgtg 600

aacattaagc tggatattac cagccataat gaagattata ccattgtgga acagtatgag 660

cgtgcggagg gacggcatag cacgggcgga atggatgaac tgtataaata a 711

<210> 16

<211> 59

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 16

gctctagagt ttttttggga attcacacac aggagagct gatggtgtct aaaggcgag 59

<210> 17

<211> 37

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 17

gctctagatt atttatacag ttcatccatt ccgcccg 37

<210> 18

<211> 744

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 18

gtttttttgg gaattcacac acaggagggag ctgatggtgt ctaaaggcga ggaagataat 60

atggcgatta tcaaagaatt tatgcgtttt aaagtgcata tggaaggcag cgtgaatggg 120

catgagtttg aaattgaagg cgaaggagaa ggccgtccgt atgaaggcac ccagaccgct 180

aaactgaaag tgaccaaagg cggaccactg ccgtttgcgt gggacattct gagcccgcag 240

tttatgtatg gcagcaaagc gtatgtgaaa catccggcgg atattccgga ttatctgaaa 300

ctgagctttc cggagggctt caaatgggaa cgtgtgatga attttgaaga tggcggcgtg 360

gtgaccgtga cccaggatag cagcctgcaa gacggcgaat tcatttacaa ggtgaagctg 420

cgtggcacca actttcccag cgatggcccg gtgatgcaga aaaagaccat gggctgggag 480

gcgagcagcg aacgtatgta cccggaggat ggcgcgctga agggcgaaat taagcagcgt 540

ctgaagttaa aagatggtgg gcactatgat gcggaagtga aaaccaccta taaagcgaaa 600

aaaccggtgc agttaccagg cgcttataat gtgaacatta agctggatat taccagccat 660

aatgaagatt ataccattgt ggaacagtat gagcgtgcgg agggacggca tagcacgggc 720

ggaatggatg aactgtataa ataa 744

<210> 19

<211> 57

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 19

gatatcgaat tcttaacttt aagaaggaat atacatatgg tgtctaaagg cgaggaa 57

<210> 20

<211> 40

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 20

aaagttaaga attcgatatc ggcactggcc gtcgttttac 40

<210> 21

<211> 42

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 21

gaattcgagc tcggtaccat cgataagctt gatatcgaat tc 42

<210> 22

<211> 41

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 22

gatggtaccg agctcgaatt cggcactggc cgtcgtttta c 41

<210> 23

<211> 51

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 23

gaattcttaa ctttaagaag gaatatacat atgaacgtta ttgcaatatt g 51

<210> 24

<211> 28

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 24

gcctctagac cacttccctt gtacgagc 28

<210> 25

<211> 35

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 25

gccaagcttg atatcgaatt ctaacttta agaag 35

<210> 26

<211> 31

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 26

ggggtaccga gtgagctgat accgctcgcc g 31

<210> 27

<211> 30

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 27

ggatcgatag ctgtttcctg tgtgaaattg 30

<210> 28

<211> 286

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 28

gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag 60

gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa 120

tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat 180

gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg 240

ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagct 286

Claims (15)

1. Logarithmic phase specificity promoter, it is characterized in that, described promoter is the nucleotide sequence shown in any one of SEQ ID NO: 1-10. 2. Contain the biological material of the described promoter of claim 1, described biological material comprises recombinant DNA, expression cassette, transposon, plasmid vector, viral vector or engineered bacterium. 3. any of the following applications of the promoter according to claim 1: 1) Application as a logarithmic phase-specific promoter in prokaryotes; 2) Application in the construction of recombinant DNA, expression cassettes, transposons, plasmid vectors, viral vectors or engineering bacteria. 4. A recombinant DNA, characterized in that, the promotor described in claim 1 is operably connected to a downstream target gene; the target gene is selected from nucleic acid encoding a protein, nucleic acid encoding a ribozyme, and encoding an antisense RNA nucleic acid. 5. The recombinant DNA according to claim 4, wherein the protein is an enzyme, a hormone, an antibody or a growth factor. 6. The recombinant DNA according to claim 5, wherein the enzyme is selected from the group consisting of oxidoreductase, transferase, hydrolase, lyase, isomerase, and ligase. 7. An expression vector, characterized in that the expression vector comprises the recombinant DNA according to any one of claims 4-6. 8. A transformant, characterized in that, the transformant is a host bacterium carrying the expression vector according to claim 7. 9. The transformant according to claim 8, wherein the host bacteria are selected from the group consisting of Escherichia , Corynebacterium , Brevibacterium , Streptomyces ), species in the genus Hafnia . 10. The transformant according to claim 8, wherein the host bacterium is selected from Escherichia coli (E.coli), Bacillus subtilis (B.subtilis), Streptomyces coelicolor ( S.coelicolor ), hive Hafnia ( H. alvei ), Corynebacterium glutamicum ( C. glutamicum ). 11. The transformant according to claim 10, characterized in that, the host bacteria is Escherichia coli ( E. coli ) or Hafnia alvei ( H. alvei ). 12. Use of the transformant according to any one of claims 8-11 in fermentative production of amino acids, polypeptides or proteins. 13. The engineering bacterium that produces lysine decarboxylase is characterized in that, described engineering bacterium contains the promoter described in claim 1, or described engineering bacterium carries the plasmid that contains lysine decarboxylase gene expression cassette, and described The lysine decarboxylase gene expression cassette is driven by the promoter described in claim 1 to express the lysine decarboxylase gene. 14. A method for fermentative production of 1,5-pentanediamine, characterized in that it comprises utilizing the engineered bacterium according to claim 13 to ferment and produce lysine decarboxylase, and then utilizing the obtained lysine decarboxylase to catalyze L- Lysine is converted to 1,5-pentanediamine. 15. The method according to claim 14, characterized in that, the engineering bacterium is fermented to obtain an enzyme fermentation liquid, and the enzyme fermentation liquid is mixed with L-lysine, L-lysine salt solution or L-lysine The fermentation broth is mixed to catalyze the transformation of L-lysine into 1,5-pentanediamine.