KR20170108880A - Novel Peptide for Enhancing Expression Efficiency and Fusion Protein Including the Same - Google Patents

Abstract

The present invention relates to a novel peptide for enhancing the expression efficiency of a target protein and a fusion protein including the novel peptide. The novel peptide according to the present invention can improve the expression efficiency of the target protein. The fusion protein having the peptide can improve the expression efficiency of the target protein as well as the water solubility to be usefully used for the production of a recombinant target protein.

Description

[0001] The present invention relates to novel peptides for enhancing the expression efficiency of a target protein, and fusion proteins comprising the novel peptides. [0002]

The present invention relates to novel peptides for enhancing the expression efficiency of a target protein and fusion proteins comprising the novel peptides. More specifically, it relates to a peptide consisting of four amino acids derived from LysRS (Lysyl tRNA synthetase) and a fusion protein comprising hRBD (human LysRS RNA binding domain) as a fusion partner with the peptide.

The key to modern biotechnology is the production of recombinant proteins, the most important of which is the production of large quantities of recombinant proteins. The recombinant proteins produced are highly important for the production and recovery of active foreign proteins, crystallization for functional studies, and industrialization. So far, many recombinant protein production studies have been conducted using E. coli, because it is easy to manipulate E. coli, has short growth time, safe expression, low cost, and can easily change scale. However, since recombinant proteins of exogenous origin produced in E. coli have low expression levels or lack the proper "post-translational chaperons" or "posttranslational processing & (Recombinant protein expression in Escherichia coli, Current Opinion in Biotechnology (1999), 10: 411-421). In addition, the recombinant protein is expressed as an insoluble protein inclusion body.

In order to solve these problems, there has been developed a method of expressing a recombinant protein by fusion of a highly water-soluble protein such as MBP, NusA, SUMO, Thioredoxin or GST (Dominic Esposito and Deb K Chatterjee, Enhancement of soluble protein expression through the use of fusion tags, Current Opinion in Biotechnology (2006), 17: 353-358). In addition, methods for increasing the acceptability of recombinant proteins using RNA binding domains such as LysRS (lysyl tRNA synthetase) have been attempted (Choi, SI, et al., Protein solubility and folding enhancement by interaction with RNA, PLoS ONE (2008) 3: e2677). However, LysRS is not only a large protein of about 100 kDa (monomer size = about 58 kDa) but also because of steric hindrance due to structural limitations of dimer formation, 3-dimensional structure formation, dimerization with more than a dimer, or formation of monomers can be prevented (Choi SI et al., Protein solubility and folding enhancement by interaction with RNA, PLoS ONE (2008), 3: e2677) .

In order to solve the problems of the prior art, the present inventors have studied the water solubility and expression efficiency of proteins, and peptides composed of four amino acids derived from Lysyl tRNA synthetase (LysRS) play a crucial role in enhancing the expression efficiency of a target protein , Confirming that a fusion protein comprising a peptide consisting of four amino acids derived from LysRS and a fusion partner comprising hRBD (human LysRS RNA binding domain) can enhance not only the expression efficiency of the target protein but also the water solubility, .

Accordingly, the object of the present invention is to provide a peptide consisting of four amino acids derived from LysRS, an expression vector containing the peptide, and a host cell transformed with the expression vector. The present invention also provides a fusion protein comprising hRBD as a fusion partner of a peptide consisting of four amino acids derived from LysRS, an expression vector comprising said fusion protein, and a host cell transformed with said expression vector The purpose.

According to a first embodiment,

The present invention provides a peptide for improving the expression efficiency of a target protein comprising the amino acid sequence of SEQ ID NO: 1.

As used herein, the term " aminoacyl tRNA synthetase "refers to an aminoacyl tRNA synthetase that binds a corresponding amino acid and a tRNA in a cell, and the thus formed aminoacyl tRNA is transferred to an elongation factor or a ribosome And is used for protein synthesis. The binding specificity of the aminoacyl tRNA synthetase to amino acids and tRNA plays an important role in maintaining the accuracy of the protein synthesis process.

As used herein, the term " lysyl tRNA synthetase (LysRS) "is a member of the aminoacyl tRNA synthetase and is used in several mammals to regulate the various functions of proteins constituting aminoacyl tRNA synthetase Form a macromolecular complex that acts as a molecular reservoir.

As used herein, the term "target protein" refers to any protein that a person skilled in the art intends to produce in large quantities and which is capable of expressing in a host cell by inserting a polynucleotide encoding the protein into a recombinant expression vector .

As used herein, the term "recombinant protein" or "fusion protein" refers to a protein that is linked to another protein at the N- or C-terminus of the original target protein sequence, .

In the peptide for enhancing the expression efficiency of a target protein according to the present invention, the amino acid sequence of SEQ ID NO: 1 may be derived from LysRS (Lysyl tRNA synthetase).

In the peptide for enhancing the expression efficiency of the target protein according to the present invention, the peptide may bind to the N-terminal of the target protein.

In the peptide for enhancing the expression efficiency of a target protein according to the present invention, the target protein may be selected from the group consisting of an antigen, an antibody, a cell receptor, an enzyme, a structural protein, a serum and a cell protein.

The present invention provides an expression vector comprising a gene sequence which binds to the N-terminus of a target protein and is composed of the amino acid sequence of SEQ ID NO: 1 and which encodes a peptide for enhancing the expression efficiency of a target protein.

In one embodiment of the present invention, the expression vector comprises a polynucleotide encoding a target protein and a polynucleotide encoding a peptide represented by SEQ ID NO: 1 bound to the 5'-terminal of the polynucleotide encoding the target protein .

As used herein, the term "expression vector" is a linear or circular DNA molecule consisting of a fragment encoding a polypeptide of interest operably linked to an additional fragment provided in the transcription of the expression vector. Such additional fragments include promoter and termination coding sequences. The expression vector also includes at least one replication origin, at least one selection marker, a polyadenylation signal, and the like. Expression vectors are generally derived from plasmid or viral DNA, or contain both elements.

The term "operably linked ", as used herein, indicates that fragments are arranged such that transcription in the promoter begins and proceeds through the coding sequence to the termination cipher.

In the expression vector according to the present invention, the expression vector may be a plasmid, a viral vector, a phage particle or a genome insert. The expression vector may be transformed into a host cell and then cloned or integrated into the genome of the host cell irrespective of the genome of the host cell.

The present invention also provides a host cell transformed with an expression vector comprising a gene sequence which binds to the N-terminus of a target protein and is composed of the amino acid sequence of SEQ ID NO: 1 and which encodes a peptide for enhancing expression efficiency of a target protein I want to.

The term "transformation" or "introduction" as used herein means that DNA is introduced into a host and the DNA becomes replicable as an extrachromosomal element or by chromosome integration completion. Methods for transforming an expression vector according to the present invention include electrophoresis, calcium phosphate (CaPO ₄ ), calcium chloride (CaCl ₂ ), microinjection, polyethylene glycol (PEG), DEAE- Tran method, cationic liposome method, or lithium acetate-DMSO method, but is not limited thereto.

In the host cell according to the present invention, the host cell is preferably a host cell having high efficiency of DNA introduction and high efficiency of expression of the introduced DNA, and all microorganisms including prokaryotic and eukaryotic cells can be used. Preferably, the host cell may be E. coli .

According to a second embodiment,

The present invention provides a fusion protein for enhancing the expression efficiency and water solubility of a target protein comprising hRBD (human LysRS RNA binding domain) as a peptide consisting of the amino acid sequence of SEQ ID NO: 1 and its fusion partner.

As used herein, the term " human LysRS RNA binding domain "refers to a native N-terminal extension region involved in the interaction between human RNA-derived RNA and other proteins it means.

In the fusion protein for enhancing the expression efficiency and water solubility of the target protein according to the present invention, the amino acid sequence of SEQ ID NO: 1 may be derived from LysRS (Lysyl tRNA synthetase).

In the fusion protein for enhancing the expression efficiency and water solubility of the target protein according to the present invention, the peptide may bind to the N-terminal of the target protein.

The fusion protein for enhancing the expression efficiency and water solubility of the target protein according to the present invention may comprise the amino acid sequence of SEQ ID NO: 3.

In the fusion protein for enhancing the expression efficiency and water solubility of a target protein according to the present invention, the target protein may be selected from the group consisting of an antigen, an antibody, a cell receptor, an enzyme, a structural protein, a serum and a cell protein have.

The present invention also relates to a method for enhancing the expression efficiency and water solubility of a target protein having a fusion protein comprising a peptide consisting of the amino acid sequence of SEQ ID NO: 1 and a polynucleotide encoding hRBD as a fusion partner thereof, Lt; / RTI >

In one embodiment of the present invention, the expression vector comprises a polynucleotide encoding a target protein and a polynucleotide encoding a fusion protein of SEQ ID NO: 5 bound to the 5'-terminal of the polynucleotide encoding the target protein And an expression vector.

In the expression vector according to the present invention, the expression vector may be a plasmid, a viral vector, a phage particle or a genome insert. The expression vector may be transformed into a host cell and then cloned or integrated into the genome of the host cell irrespective of the genome of the host cell.

The present invention also relates to a method for enhancing the expression efficiency and water solubility of a target protein having a fusion protein comprising a peptide consisting of the amino acid sequence of SEQ ID NO: 1 and a polynucleotide encoding hRBD as a fusion partner thereof, The present invention provides a host cell transformed with an expression vector for < RTI ID = 0.0 >

In the host cell according to the present invention, the host cell is preferably a host cell having high efficiency of DNA introduction and high efficiency of expression of the introduced DNA, and all microorganisms including prokaryotic and eukaryotic cells can be used. Preferably, the host cell may be E. coli .

The present invention also provides a method for producing a target protein with enhanced expression efficiency. The method for producing the target protein comprises:

(A) preparing an expression vector comprising a polynucleotide encoding a target protein and a polynucleotide encoding a peptide that promotes the expression efficiency of a target protein bound to the 5'-terminal of the polynucleotide encoding the target protein ;

(B) introducing the expression vector into a host cell to prepare a transformant; and

(C) culturing the transformant to induce the expression of the recombinant target protein, and obtaining the same.

In one embodiment of the present invention, the peptide that enhances the expression efficiency of the target protein may be derived from LysRS (Lysyl tRNA systhetase), preferably the amino acid sequence shown in SEQ ID NO: 1.

In another embodiment of the present invention, the peptide for enhancing the expression efficiency of the target protein may be a fusion protein comprising a peptide sequence derived from LysRS (Lysyl tRNA systhetase) and hRBD, Lt; / RTI >

The peptide consisting of four amino acids derived from LysRS according to the present invention can improve the expression efficiency of a target protein and the fusion protein containing hRBD as a fusion partner with the peptide can be used not only for the expression efficiency of a target protein, , It can be usefully used for the production of a recombinant target protein.

1 is a graph showing the effect of the peptide of the present invention according to Example 1 on the expression efficiency of EGFP-HBx protein.
FIG. 2 is a graph showing the effect of the peptides of the present invention and the fusion protein comprising them according to Examples 2 and 3 on the expression efficiency and water solubility of CsTA381 and CsTA1953 proteins.
FIG. 3 shows the results of SDS-PAGE showing the effect of the peptide of the present invention and the fusion protein comprising it according to Example 4 on the expression efficiency and water solubility of CsTA37 and CsTA422 proteins.
FIG. 4 shows experimental results comparing the effect of increasing the expression efficiency of a target protein in the peptide sequence according to the present invention compared to other peptide sequences.

Hereinafter, various embodiments are provided to facilitate understanding of the present invention. The following examples are provided to facilitate understanding of the invention and are not intended to limit the scope of the invention.

<Experimental Method>

1. Preparation of protein expression vector

Protein solubility and folding enhancement by interaction with RNA, PLoS ONE (2008), 3: e2677) were used as expression vectors for pGE-LysRS. The expression of pGE-LysRS is regulated by the T7 promoter and the LysRS gene is cut out using one of the cleavage sites in Nde1 and MCS (Kpn1-BamH1-EcoRV-Sal1-Hind3) and EGFP-HBx or hRBD , Or other proteins. At this time, the amino acid sequence of SEQ ID NO: 1 was inserted at the N-terminal position of the inserted protein. Then, the target proteins to be expressed were inserted using two produced enzyme sites in the MCS of the produced hRBD or the amino acid-containing hRBD vector of SEQ ID NO: 1.

2. Protein expression and SDS-PAGE

The prepared protein expression vector was transformed into BL21 (DE3) or BL21 * (DE3) -pLysS or BL21 (DE3) -pLysE competent cells. All transformed Escherichia coli were cultured in LB medium containing 50 μg / ml of ampicillin. Escherichia coli transformed into BL21 * (DE3) -pLysS or BL21 (DE3) -pLysE contained 34 μg / ml of chloramphenicol Were added to the medium. The incubation temperature varies from protein to protein and was cultured at 25 to 37 ° C. When the OD ₆₀₀ value of E. coli is 0.5 or more, IPTG is added at a level of 0 μM to 1 mM in order to activate the T7 promoter. For the sufficient production of the protein, IPTG is added for 3 hours at 37 ° C. or 25 ℃ for 5 hours. The well-cultured E. coli was centrifuged and the supernatant was removed and stored. Then, 0.3 ml of PBS was added to 5 ml of the LB medium to produce E. coli, and lysate was prepared by sonication. The total lysate, soluble fraction and pellet fraction of the lysate were separated by SDS-PAGE and analyzed by SDS-PAGE. The lysates were separated by centrifugation and then separated into soluble fraction and pellet fraction. Respectively.

3. Western Blotting (Western blotting)

The sample to be analyzed was subjected to SDS-PAGE and immediately transferred to a PVDF membrane. The voids of the PVDF membrane, which were not filled with the sample protein, were blocked with 5% skim milk in TBST (pH 7.5 Tris-buffered saline, + 0.05% tween 20). The membranes were then washed three times with TBST and then treated with Anti-his tag (1: 20000) (QIAGEN, 34660) for 16 hours at 4 ° C. The membranes were washed again with TBST three times and then treated with HRP-conjugated anti-mouse antibody (Sigma, A4416) as a secondary antibody for 1 hour at room temperature and again with TBST for 3 times After washing, WEST-ZOL (Intron, Korea) was treated and confirmed as a film.

< Example >

Example  1. According to the invention The peptide EGFP - HBx  Effect on protein expression efficiency

(1) SDS-PAGE analysis

The LysRS was removed from the pGE LysRS plasmid using Nde1 and Hind3 restriction enzymes and the gene sequence (SEQ ID NO: 2) coding for the EGFP-HBx gene direct sequence and SEQ ID NO: 1 was added to the N- Two fusion forms were inserted. The vector is expressed by the T7 promoter, which is regulated by the lac operator and regulated by IPTG. Two recombinant plasmids were transformed into BL21 (DE3) -pLysE competent cells, respectively, and expressed at 37 ° C for 3 hours. At this time, IPTG was treated at four concentrations of 0, 10, 20 and 40 μM. As a result, it was confirmed that the fusion form of the EGFP-HBx gene was expressed better than the direct form (FIG. 1A).

(2) Western blotting analysis

Since the proteins of Escherichia coli itself form a background, accurate quantification of the total fraction is difficult with SDS-PAGE results. Western blotting (WB) was performed, and the WB result was quantitated using BioD (S / W) for analysis. As a result, it was confirmed that the fusion form of the EGFP-HBx gene increased the expression level at least twice as compared to the direct form (FIG. 1B).

 On the other hand, the EGFP-HBx protein is almost completely absent in the soluble fraction, and most of it is found in the pellet fraction. Since the pellet fraction contains no Escherichia coli proteins, background is much smaller than the total fraction. Therefore, the quantification using the pellet fraction and the comparison with the IPTG 10 μM (orange bar) before saturation showed that the fusion form of the EGFP-HBx gene had an expression level at least 7 times higher than that of the direct form (Fig. 1C).

Example  2. According to the present invention The peptide CsTA381  And CsTA1953  Effect on protein expression efficiency

In order to examine the effect of peptides according to the present invention on the expression of CsTA381 and CsTA1953 proteins, which are known to be poorly expressed in Escherichia coli, the direct and fusion forms of the two proteins were expressed as pGE LysRS plasmid. A total of four recombinant plasmids were transformed into BL21 * (DE3) -pLysS competent cells and expressed at 37 ° C. At this time, IPTG was treated at a concentration of 1 mM and cultured for 3 hours. As a result, the fusion forms of the CsTA381 and CsTA1953 proteins were found to have an increased expression level compared to the direct form (FIGS. 2A and 2B).

Example  3. According to the present invention Peptides  And hRBD  Lt; RTI ID = 0.0 &gt; CsTA381 &lt; / RTI & CsTA1953  Effect on protein expression and water solubility

HRBD was used as a fusion partner to increase the water solubility of CsTA381 and CsTA1953 proteins according to Example 2. The hRBD is represented by the amino acid sequence of SEQ ID NO: 3, and the fusion protein (mseq-hRBD) obtained by binding the peptide sequence of SEQ ID NO: 1 to hRBD is represented by the amino acid sequence of SEQ ID NO:

LysRS was cut out using Nde1 and Kpn1 in the pGE LysRS plasmid, and the DNAs of SEQ ID NO: 4 and SEQ ID NO: 6, which are gene sequences coding for hRBD and mseq-hRBD, were inserted at the positions, respectively. Then, CsTA381 gene and CsTA1953 gene were inserted into pGE hRBD plasmid and pGE mseq-hRBD plasmid using BamH1 and Hind3, respectively.

As a result, it was not confirmed whether the 5 'hRBD binding form of the CsTA381 and CsTA1953 proteins improved the water solubility due to poor expression efficiency. However, mseq-hRBD binding (CsTA381 and CsTA1953) proteins were improved (Fig. 2C and 2D). In addition, the protein bands of hRBD binding form and mseq-hRBD binding form were measured and compared by densitometric scanning. As a result, the expression level of mseq-hRBD binding form of CsTA381 and CsTA1953 protein was at least 4 times (Figs. 2E and 2F). In the mseq-hRBD binding form, the level of expression was much higher than that of the binding form of SEQ ID NO: 1, confirming that there is a synergistic effect of hybrid type. Therefore, when the mseq (SEQ ID NO: 1) sequence and the hRBD (SEQ ID NO: 2) sequence are used as a hybrid form (amino acid sequence of mseq-hRBD), not only the water solubility of the objective protein but also the expression efficiency can be greatly increased.

Example  4. According to the present invention Peptides  And hRBD  Lt; RTI ID = 0.0 &gt; CsTA37 &lt; CsTA422  Effect on protein expression and water solubility

In order to confirm the effect of mseq-hRBD (SEQ ID NO: 5) binding form confirmed in Examples 2 and 3 in other proteins, CsTA37 and CsTA422 were also applied to two proteins. The direct form and mseq-hRBD (SEQ ID NO: 5) binding form plasmids were prepared in the same manner as in Example 3, and expression was carried out at 30 ° C in BL21 E. coli. As a result, it was confirmed that the direct form showed almost no expression and the water solubility was poor, but it was confirmed that the mseq-hRBD-binding form was expressed well in a water-soluble form (FIG. 3).

Example  5. According to the present invention Of peptide  Expression Increase efficiency  Compare effects

In order to confirm the effect of the mseq (SEQ ID NO: 1) peptide sequence according to the present invention in increasing the expression efficiency of a target protein, the peptide sequence of SEQ ID NO: 7 was compared with another peptide sequence derived from LysRS using the same method as in Example 1 above.

Specifically, LysRS was removed from the pGE LysRS plasmid using Nde1 and Hind3 restriction enzymes and a gene sequence (SEQ ID NO: 2, fusion form (SEQ ID NO: 2) coding for the peptide added at the N-terminal of SEQ ID NO: ; Mseq) and a gene sequence (SEQ ID NO: 8, fusing form; Mseqhaq) coding for a peptide added at the N-terminus of SEQ ID NO: 7. The vector is expressed by the T7 promoter, which is regulated by the lac operator and regulated by IPTG. The three recombinant plasmids were transformed into BL21 (DE3) -pLysE competent cells, respectively, and proteins were expressed at 37 ° C for 3 hours. At this time, IPTG was treated at four concentrations of 0, 10, 20 and 40 μM.

As a result, it was confirmed that when the expression of the GFP gene was expressed together with the mseq (SEQ ID NO: 1) peptide sequence according to the present invention, it was increased by 27% as compared with when it was expressed together with the peptide sequence of Mseqhaq (SEQ ID NO: 7) 4).

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

<110> Industry-Academic Cooperation Foundation. Yonsei University <120> Novel Peptide for Enhancing Expression Efficiency and Fusion          Protein Including the Same <130> 1062255 <150> KR 10-2016-0032451 <151> 2016-03-18 <160> 8 <170> KoPatentin 3.0 <210> 1 <211> 4 <212> PRT <213> Artificial Sequence <220> <223> MSEQ Protein seq <400> 1 Met Ser Glu Gln   One <210> 2 <211> 12 <212> DNA <213> Artificial Sequence <220> <223> MSEQ DNA seq <400> 2 atgtctgaac aa 12 <210> 3 <211> 74 <212> PRT <213> Artificial Sequence <220> <223> hRBD Protein seq <400> 3 Met His Ala Gln Ala Ala Val Gln Ala Ala Glu Val Lys Val Asp Gly   1 5 10 15 Ser Glu Pro Lys Leu Ser Lys Asn Glu Leu Lys Arg Arg Leu Lys Ala              20 25 30 Glu Lys Lys Val Ala Glu Lys Glu Ala Lys Gln Lys Glu Leu Ser Glu          35 40 45 Lys Gln Leu Ser Gln Ala Thr Ala Ala Ala Thr Asn His Thr Thr Asp      50 55 60 Asn Gly Val Gly Pro Glu Glu Glu Ser Val  65 70 <210> 4 <211> 222 <212> DNA <213> Artificial Sequence <220> <223> hRBD DNA seq <400> 4 atgcacgcac aggcggccgt gcaggcggcc gaggtgaaag tggatggcag cgagccgaaa 60 ctgagcaaga atgagctgaa gagacgcctg aaagctgaga agaaagtagc agagaaggag 120 gccaaacaga aagagctcag tgagaaacag ctaagccaag ccactgctgc tgccaccaac 180 cacaccactg ataatggtgt gggtcctgag gaagagagcg tg 222 <210> 5 <211> 77 <212> PRT <213> Artificial Sequence <220> <223> Fusion protein Protein seq <400> 5 Met Ser Glu Gln His Ala Gln Ala Ala Val Gln Ala Ala Glu Val Lys   1 5 10 15 Val Asp Gly Ser Glu Pro Lys Leu Ser Lys Asn Glu Leu Lys Arg Arg              20 25 30 Leu Lys Ala Glu Lys Lys Val Ala Glu Lys Glu Ala Lys Gln Lys Glu          35 40 45 Leu Ser Glu Lys Gln Leu Ser Gln Ala Thr Ala Ala Ala Thr Asn His      50 55 60 Thr Asp Asn Gly Val Gly Pro Glu Glu Glu Ser Val  65 70 75 <210> 6 <211> 231 <212> DNA <213> Artificial Sequence <220> <223> Fusion protein DNA seq <400> 6 atgtctgaac aacacgcaca ggcggccgtg caggcggccg aggtgaaagt ggatggcagc 60 gagccgaaac tgagcaagaa tgagctgaag agacgcctga aagctgagaa gaaagtagca 120 gagaaggagg ccaaacagaa agagctcagt gagaaacagc taagccaagc cactgctgct 180 gccaccaacc acaccactga taatggtgtg ggtcctgagg aagagagcgt g 231 <210> 7 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> MSEQHAQ Protein seq <400> 7 Met Ser Glu Gln His Ala Gln   1 5 <210> 8 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> MSEQHAQ DNA seq <400> 8 atgtctgaac aacacgcaca g 21

Claims (19)

Wherein the peptide is bound to the N-terminus of the target protein and is composed of the amino acid sequence of SEQ ID NO: 1.
The method according to claim 1,
Wherein the amino acid sequence of SEQ ID NO: 1 is derived from LysRS (Lysyl tRNA systhetase).
The method according to claim 1,
Wherein the target protein is at least one selected from the group consisting of an antigen, an antibody, a cell receptor, an enzyme, a structural protein, a serum and a cell protein.
A polynucleotide encoding a target protein; And
A polynucleotide encoding a peptide according to any one of claims 1 to 3 linked to the 5'-end of a polynucleotide encoding said protein of interest;
&Lt; / RTI &gt;
A host cell transformed with an expression vector according to claim 4.
6. The method of claim 5,
Wherein said host cell is E. coli .
A peptide which is bound to the N-terminus of the target protein and consists of the amino acid sequence of SEQ ID NO: 1; And
Wherein the fusion protein comprises hRBD (human LysRS RNA binding domain) as a fusion partner of the peptide.
8. The method of claim 7,
Wherein the hRBD comprises the amino acid sequence of SEQ ID NO: 3.
8. The method of claim 7,
Wherein the target protein is selected from the group consisting of an antigen, an antibody, a cell receptor, an enzyme, a structural protein, a serum and a cell protein.
8. The method of claim 7,
Wherein the fusion protein comprises the amino acid sequence of SEQ ID NO: 5.
A polynucleotide encoding a target protein; And
A polynucleotide encoding a fusion protein according to any one of claims 7 to 10 linked to the 5'-end of a polynucleotide encoding said protein of interest;
&Lt; / RTI &gt;
12. A host cell transformed with an expression vector according to claim 11.
13. The method of claim 12,
Wherein said host cell is E. coli .
(A) preparing an expression vector comprising a polynucleotide encoding a target protein and a polynucleotide encoding a peptide that enhances the expression efficiency of a target protein bound to the 5'-terminal of the polynucleotide encoding the target protein ;
(B) introducing the expression vector into a host cell to prepare a transformant; and
(C) culturing the transformant to induce expression of the recombinant target protein, and obtaining the same.
15. The method of claim 14,
Wherein the peptide for enhancing the expression efficiency of the target protein is derived from LysRS (Lysyl tRNA systhetase).
15. The method of claim 14,
Wherein the peptide enhancing expression efficiency of the target protein is composed of the amino acid sequence represented by SEQ ID NO: 1.
15. The method of claim 14,
Wherein the peptide for enhancing the expression efficiency of the target protein is a fusion protein comprising a peptide sequence derived from LysRS (Lysyl tRNA systhetase) and hRBD.
18. The method of claim 17,
Wherein the hRBD comprises an amino acid sequence represented by SEQ ID NO: 3.
18. The method of claim 17,
Wherein the fusion protein comprises an amino acid sequence represented by SEQ ID NO: 5.

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