KR101494534B1 - Novel isolated peptide for binding to somatic structure and uses thereof - Google Patents

Abstract

The present invention relates to a novel isolated peptide comprising the amino acid sequence of SEQ ID NO: 1 or 2, a polynucleotide encoding the peptide, an expression vector comprising the polynucleotide, a transformant into which the expression vector is introduced into a host cell, A method for producing the peptide using the transformant, a crosslinking agent comprising the peptide, and a composition comprising the crosslinking agent. The peptides provided in the present invention can be universally bound to biological structures such as hair, skin, and teeth, and can be used as a crosslinking agent for various pharmaceutical compositions, quasi-drug compositions, and cosmetic compositions. Therefore, Compositions for quasi-drugs, and cosmetic compositions.

Description

[0001] The present invention relates to novel isolated peptides that bind to living body structures,

The present invention relates to a novel isolated peptide which binds to a biological structure and its use, and more particularly to a novel isolated peptide comprising the amino acid sequence of SEQ ID NO: 1 or 2, a polynucleotide encoding said peptide , An expression vector comprising the polynucleotide, a transformant into which the expression vector is introduced, a method for producing the peptide using the transformant, a crosslinking agent comprising the peptide, and a composition comprising the crosslinking agent .

Generally, the pharmaceutical composition for preventing or treating various diseases occurring in a living body is prepared in various formulations in consideration of the symptoms of the disease, the condition of the patient, the condition of the tissue caused by the disease, and the like. Despite the fact that various formulations have been developed to date and the active ingredient is effectively administered to the disease-induced tissue, there is a problem in that the utilization ratio of the effective ingredient is only low, which effectively transmits the effective ingredient to the disease- It is because it can not be. As means for solving such problems, a method for improving the in vivo residual ratio of the active ingredient has been studied, and as a result, a formulation having improved in vivo persistence such as a sustained release preparation has been developed. However, since such a sustained-release preparation acts in such a manner that an excessive amount of the active ingredient is administered in the body and gradually releases it, it is excellent in convenience of decreasing the administration frequency, but the released active ingredient is delivered to the disease- Is not improved, it can not be said to be effective in terms of improving the in vivo residual ratio of the active ingredient.

As one of the measures to fundamentally solve the above problem, there has been sought a method for increasing the residual rate in vivo by binding the active ingredient itself to the surface of the disease-induced vital structure, and for this purpose, There have been a lot of researches to develop cross-linking agents. As a result, studies have been conducted to bind an active ingredient to a disease-induced tissue using an antibody or the like as a cross-linking agent, and some of the drugs thus developed have been commercialized. However, when the antibody is used as a cross-linking agent, it is specifically bound to a specific tissue, resulting in an increase in production cost, which inevitably leads to an increase in the price of the therapeutic agent.

In the case of pharmaceutical compositions, such as anticancer agents, which are likely to cause serious side effects in tissues other than the diseased tissue, a cross-linking agent, such as an antibody capable of specifically binding to the diseased tissue, Must be used. However, in the case of a pharmaceutical composition that does not cause serious side effects in normal tissues, a cross-linking agent capable of binding to a disease-causing tissue in general can be used rather than a cross-linking agent such as an antibody capable of specifically binding to the diseased tissue It is desirable to use a cross-linking agent that can be used in general use as such, but no research results have been reported so far.

Under these circumstances, the inventors of the present invention have extensively studied to develop a crosslinking agent capable of binding to a living body structure or disease-causing tissue in general, and as a result, it has been found that a peptide comprising the amino acid sequence of SEQ ID NO: The present invention has been completed based on this finding.

It is an object of the present invention to provide novel isolated peptides.

It is another object of the present invention to provide a polynucleotide encoding said peptide.

It is still another object of the present invention to provide an expression vector comprising the polynucleotide.

It is still another object of the present invention to provide a transformant into which the above expression vector has been introduced.

It is still another object of the present invention to provide a method for producing the peptide using the transformant.

It is still another object of the present invention to provide a crosslinking agent comprising the peptide.

It is another object of the present invention to provide a composition comprising the crosslinking agent.

In one embodiment, the present invention provides a novel isolated peptide comprising the amino acid sequence of SEQ ID NO: 1 or 2 and exhibiting bio-structure binding activity.

SEQ ID NO: 1: KVWQLQLPQWIN

SEQ ID NO: 2: KVWTLSNDYIEF

The inventors of the present invention focused on the fact that most important biological structures are related to proteins while carrying out various studies to develop a method of retaining a useful component in a biological structure for a long period of time. That is, since the hair belonging to the biological structure is composed of a cuticle layer including a keratin protein on the surface, the skin belonging to the biological structure is composed of keratin protein and lipid, and the teeth belonging to the biological structure are composed of collagen and inorganic substances, It has been expected that the use of the protein or peptide capable of effectively binding with the protein or other components contained in the above-described biological structure will allow the useful component to remain in the biological structure for a long period of time. In order to develop peptides capable of performing a cross-linking role between the bio-structures and useful components, peptides showing ability to bind to various biological structures such as hair, skin, and teeth were selected using a phage display method. It was confirmed that the selected peptides had excellent binding to hair and excellent binding to skin surface and tooth surface.

Therefore, it is understood that the novel peptide comprising the amino acid sequence of SEQ ID NO: 1 or 2 provided in the present invention can be used as a crosslinking agent contained in a composition such as a pharmaceutical composition, a quasi-drug composition, a cosmetic composition, there was.

The peptide provided by the present invention includes the amino acid sequence of SEQ ID NO: 1 or 2, and may include various amino acids at the N-terminal or C-terminal of the amino acid sequence of SEQ ID NO: 1 or 2, And all peptides to which the sequence has been added.

The amino acids referred to herein are abbreviated as follows according to the IUPAC-IUB nomenclature.

Alanine: A, arginine: R, asparagine: N,

Aspartic acid: D, cysteine: C, glutamic acid: E,

Glutamine: Q, Glycine: G, Histidine: H,

Isoleucine: I, leucine: L, lysine: K,

Methionine: M, phenylalanine: F, proline: P,

Serine: S, threonine: T, tryptophan: W,

Tyrosine: Y, Valine: V

The term "peptide " of the present invention means a polymer composed of an amino acid linked by an amide bond (or a peptide bond). The method for producing the peptide is not particularly limited. Preferably, the peptide is produced by using a gene recombination and protein expression system, synthesized in vitro through chemical synthesis such as peptide synthesis, a cell-free protein synthesis method, A method using an acid generator, a method using a synthesizer, and the like.

In the present invention, the peptide may be a peptide comprising the amino acid sequence of SEQ ID NO: 1 or 2, which exhibits a binding activity to a living body structure and can function as a bridge between the living body structure and the useful substance. In addition, if the amino acid sequence of the amino acid sequence of SEQ ID NO: 1 or 2 is mutated by a method such as addition, substitution, deletion or the like, the peptide may be included in the scope of the peptide of the present invention have. In addition, the peptides of the present invention may further comprise a targeting sequence, a tag, a labeled residue, a half-life or an amino acid sequence designed for a specific purpose to increase the stability of the peptide.

The term "biological structure" of the present invention means a constituent component structurally constituting a living body. For example, hair, skin, teeth, and the like.

According to one embodiment of the present invention, peptides displaying binding properties to hair are screened using a phage display method to select peptides containing the amino acid sequence of SEQ ID NO: 1 or 2 (Example 1) The binding activity of the peptides to the vital structures was measured. As a result, the peptide showed strong binding activity to the hair (Table 1), skin (Table 2) and teeth (Table 3), and the peptide of SEQ ID NO: It was confirmed that the binding activity was relatively good.

As another embodiment for achieving the above-mentioned object, the present invention provides a polynucleotide comprising a nucleotide sequence encoding the peptide.

In the present invention, the nucleotide sequence constituting the polynucleotide may be a nucleotide sequence capable of encoding the amino acid sequence of SEQ ID NO: 1 or 2 or a nucleotide sequence which is added at the N-terminus or C-terminus of the amino acid sequence of SEQ ID NO: May be a form in which a base sequence encoding various amino acid sequences that can be obtained is added to the 5'-terminal or 3'-terminal of a base sequence capable of encoding the amino acid sequence of SEQ ID NO: 1 or 2, A nucleotide sequence of SEQ ID NO: 3 or 4 capable of encoding the amino acid sequence of SEQ ID NO: 1 or 2 or a variety of amino acid sequences which may be added to the N-terminal or C-terminal of the amino acid sequence of SEQ ID NO: Or 3 ' -terminal of the nucleotide sequence of SEQ ID NO: 3 or 4 above.

SEQ ID NO: 3: 5'-AAGGTTTGGCAGCTGCAGCTGCCGCAGTGGATTAAT-3 '

SEQ ID NO: 4: 5'-TAAGGTGTGGACGCTGTCGAATGATTATATTGAGTTT-3 '

Also, as long as it is capable of coding a peptide capable of exhibiting binding activity with respect to the above-described biological structure, a polynucleotide including a base sequence having homology with the above base sequence may be included in the category of the polynucleotide provided in the present invention , Preferably 80% or more homology, and more preferably 90% or more homology. Most preferably, the polynucleotide may be a polynucleotide comprising a nucleotide sequence having a homology of at least 90% Or a polynucleotide comprising a nucleotide sequence having 95% or more homology.

The polynucleotide can be obtained from a library using phage display technology. Phage display technology is a technique for expressing a random polynucleotide on a phage surface, which comprises the steps of: 1) introducing a random sequence of oligonucleotides or gene libraries into the gene region corresponding to the coat protein pIII N- ; 2) expressing a fusion protein of a portion of the native type of coat protein and a polypeptide encoded by the oligonucleotide or gene library of the random sequence; 3) treating a receptor material capable of binding to the oligonucleotide or polypeptide encoded by the gene library; 4) eluting the phage particles bound to the receptor using low pH or competitive binding molecules; 5) amplifying the phage eluted by panning in the host cell; 6) repeating the method to obtain the desired amount; And 7) determining the amino acid sequence of the active peptide or protein from the DNA sequence of the phage clones selected by panning.

In addition, polynucleotide sequences encoding the peptides of the present invention can also be obtained using known polynucleotide synthesis methods.

In another embodiment for achieving the above object, the present invention provides an expression vector comprising the polynucleotide.

The term "expression vector" of the present invention means a gene construct comprising a gene insert that is capable of expressing a desired protein in an appropriate host cell, and an essential regulatory element operably linked to the expression of said gene insert.

In the present invention, the expression vector is not particularly limited as long as it is capable of producing the peptide by expressing the polynucleotide. Examples of the expression vector include a mammalian cell (such as a human, a monkey, a rabbit, a rat, a hamster, May be a vector capable of replicating and / or expressing the polynucleotide in eukaryotic or prokaryotic cells, including cells, yeast cells, insect cells or bacterial cells (e. G., E. coli, etc.) (PYG601BR322, pBR325, pUC118 and pUC119), Bacillus subtilis (pUC119) and pUC119 (bacillus subtilis) are operably linked to appropriate promoters so that the polynucleotides can be expressed, and can be vectors containing at least one selection marker. More preferably, Bacillus subtilis-derived plasmids (pUB110 and pTP5), yeast-derived plasmids (YEp13, YEp24 And YCp50), lambda-phage (Charon4A, Charon21A, EMBL3, EMBL4, lambda gt10, lambda gt11 and lambda ZAP), retrovirus, adenovirus, vaccinia virus, baculovirus .

As another embodiment for accomplishing the above object, the present invention provides a transformant wherein the expression vector is introduced into a host cell.

The host cell to which the expression vector provided in the present invention can be introduced is not particularly limited as long as it can produce the peptide by expressing the polynucleotide. Preferably, the host cell is a bacterial cell such as Escherichia coli, Streptomyces, Salmonella typhimurium, etc. ; Yeast cells; Fungal cells such as Pichia pastoris; Insect cells such as Drosophila and Spodoptera Sf9 cells; Animal cells such as CHO, COS, NSO, 293, Bowmanella cells; Or plant cells, and more preferably mammalian animal cells.

As another embodiment for achieving the above object, the present invention provides a method for producing the peptide using the transformant.

Specifically, a method of producing a peptide of the present invention comprises the steps of: (a) obtaining a polynucleotide encoding the amino acid sequence of SEQ ID NO: 1 or 2; (b) cloning the obtained polynucleotide to obtain an expression vector; (c) introducing the obtained expression vector into a host cell to obtain a transformant; And (d) culturing the transformant, and recovering a peptide comprising the amino acid sequence of SEQ ID NO: 1 or 2 from the transformant.

As another embodiment for achieving the above-mentioned object, the present invention provides a cross-linking agent comprising the peptide.

The peptides provided in the present invention exhibit a strong binding activity to biological structures such as hair, skin, and teeth, and can perform a crosslinking function of binding various useful components to the biological structures. Thus, the peptides provided in the present invention Can be used as a crosslinking agent to improve the usability of an active ingredient by providing a bond to a living body structure by being linked to an active ingredient of various compositions such as a pharmaceutical composition, a quasi-drug composition, and a cosmetic composition.

According to one embodiment of the present invention, each GFP-fusion peptide in which a peptide comprising the amino acid sequence of SEQ ID NO: 1 or 2 is bound to the N-terminus of the GFP protein is prepared (Examples 3-1 to 3 -3). The binding activity of these GFP-fusion peptides to the vital structures was measured. As a result, they showed strong binding activity to the hair (Table 4), skin (Table 5) and teeth (Table 6) It was confirmed that the GFP-fusion peptide containing the peptide exhibits relatively better binding activity than the GFP-fusion peptide including the peptide of SEQ ID NO: 2.

Therefore, it was found that the peptide of SEQ ID NO: 1 or 2 provided in the present invention shows activity as a cross-linking agent capable of binding a useful substance to a biological structure in general.

The peptides provided in the present invention generally exhibit binding to biological structures such as skin and teeth, and do not exhibit specific binding activity to specific biological structures. Therefore, peptides that are delivered to a target using a digestive or circulatory system It is preferable to utilize it as a cross-linking agent for the active ingredient of the pharmaceutical composition which directly adds to the target rather than the composition. For example, the peptide of the present invention can be used as a cross-linking agent for an active ingredient of a skin external preparation such as a liquid preparation, an ointment preparation, a lotion preparation, a spray, a patch, a cream, a powder, a suspension, a gel or an aerosol.

In addition, the peptide of the present invention can be used as a crosslinking agent included in oral preparations. When an oral preparation for treating the above-mentioned diseases is applied to the affected part of the oral cavity to treat an oral disease, the applied oral preparation may be diluted or removed due to the saliva continuously secreted from the oral cavity. The oral preparation containing the peptide of the present invention as a crosslinking agent can strongly bind its active ingredient to the affected part of the oral cavity to prevent dilution or removal due to saliva.

The peptides provided in the present invention exhibit a strong binding activity to biological structures such as hair, skin, and teeth, and can perform a crosslinking function of binding various useful components to the biological structures. Thus, the peptides provided in the present invention Can be used as a crosslinking agent to be added to the quasi-drug composition.

The term "quasi-drug composition" of the present invention is abbreviated to quasi-drug. The term " quasi-drug composition " refers to a drug or a drug that is used to treat or alleviate a disease of a human or an animal, Means that the action is weak or does not act directly on the human body and is similar to that which is not an apparatus or a machine "or" an agent used for sterilization, insecticide and similar purposes for the prevention of infectious diseases ".

In the present invention, since the peptide of the present invention can be universally bound to skin, teeth, hair, etc., the peptide can be used as a crosslinking agent for an active ingredient of a quasi-drug composition which can be used for skin, teeth, hair and the like. At this time, the quasi-drug composition that can be used for the skin, teeth, hair and the like is not particularly limited, but preferably includes hand wash, gagrin, disinfectant, shower foam, wet tissue, detergent soap, mild detergent, toothpaste, And the like.

The peptides provided in the present invention not only exhibit general binding activity to sites such as the skin where the cosmetic composition can be applied, but also can function as a bridge for binding various components to the skin. Thus, Can be used as a cross-linking agent for the active ingredient of the cosmetic composition.

In the present invention, the cosmetic composition to which the peptide may be incorporated as a crosslinking agent is not particularly limited, but may be a cream, an essence, a lotion, a pack, a nutritional cream, a massage cream, a cleansing cream, a cleansing foam, a cleansing water, Such as soap, liquid cleanser, bath salts, sunscreen cream, sun oil, foundation, makeup base, lipstick, eye syringe, eyeliner, mascara, eyebrow pencil, shampoo, rinse, hair treatment, hair mousse, hair liquid, Hair color preparations, hair blot preparations, color rinses, hair tonics, scallops treatments, and the like.

In another embodiment for achieving the above object, the present invention provides a composition comprising the peptide as a cross-linking agent for an active ingredient. At this time, the composition may be a pharmaceutical composition, a quasi-drug composition or a cosmetic composition.

The peptide of the present invention can be used as a cross-linking agent for an active ingredient of a pharmaceutical composition such as external preparation for skin, injectable drug for skin and oral preparation owing to its binding activity. Therefore, the peptide can be used as a external preparation for skin, Pharmaceutical compositions such as formulations may be included within the scope of the present invention. In this case, the external preparation for skin may be formulated in the form of an external preparation or a suppository, and the skin injections may be formulated in the form of sterile injectable solutions. The oral preparations may be formulated into powders, granules, tablets, capsules, Syrup, aerosol, and the like. In addition, the pharmaceutical composition may further comprise an appropriate carrier, excipient or diluent conventionally used in the production thereof. The carrier, excipient or diluent may be a filler, an extender, a binder, a wetting agent, a disintegrant, a surfactant, a lubricant, a wetting agent, a sweetener, a fragrance or a preservative. Examples of the carrier include lubricants, dextrose, sucrose, The present invention relates to a pharmaceutical composition for oral administration which comprises at least one selected from the group consisting of sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin calcium phosphate calcium silicate calcium carbonate cellulose methylcellulose, polyvinylpyrrolidone, , Propylhydroxybenzoate, talc, magnesium stearate, mineral oil, gelatin, liquid paraffin, propylene glycol, polyethylene glycol, olive oil, ethyl oleate, witepsol, macrogol, tween 61, cacao butter, laurin, Latin and the like can be used.

The peptide of the present invention can be used as a cross-linking agent for an active ingredient of a quasi-drug composition that can be used for skin, teeth, hair and the like owing to its binding activity. Therefore, the peptide can be used as a cross-linking agent and can be used for skin, teeth, Compositions may also be included within the scope of the present invention.

Because the peptide of the present invention can be used as a crosslinking agent for an active ingredient of a cosmetic composition that can be used for skin due to its binding activity, a cosmetic composition containing the peptide as a crosslinking agent can also be included in the scope of the present invention. At this time, the cosmetic composition may contain additives other than the peptide of the present invention, which are conventionally used according to the use thereof. The additives include, but are not limited to, fragrances, coloring matters, bactericides, antioxidants, preservatives, moisturizers, , Inorganic salts, synthetic high molecular materials, oils, water, surfactants, alcohols, chelating agents, etc. may be used alone or in combination.

Meanwhile, the content of the peptide of the present invention contained in the pharmaceutical composition, quasi-drug composition or cosmetic composition can be easily selected by those skilled in the art depending on their use and formulation.

The peptides provided in the present invention can be universally bound to biological structures such as hair, skin, and teeth, and can be used as a crosslinking agent for various pharmaceutical compositions, quasi-drug compositions, and cosmetic compositions. Therefore, Compositions for quasi-drugs, and cosmetic compositions.

Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

Example  1: Binding to vital structures Of peptide  Selection

In order to screen peptides binding to a living body structure, a phage display method was performed on a hair of a living body structure.

Specifically, 50 mg of hair of a healthy adult was minced and immersed in a reaction buffer (50 mM Tris pH 7.5, 150 mM NaCl, 1% BSA), and a random polynucleotide library was introduced to constantly add phage expressing various polypeptides to the surface . After the reaction was completed, the hair was washed and the phage was eluted by adding 0.2 N HCl solution, and the eluted phage was amplified. Then, the amplified phage was applied to the hair, the hair was washed, and the process of eluting the phage from the hair was performed twice.

At this time, a random polynucleotide library was used with Ph.D.-12 phage library kit (New England Biolab) and 10 ⁹ phages were added to the hair. The amplification of the phage was carried out by inoculating the extracted phage into E. coli ER2738 (New England Biolab). Specific methods are as follows: Inoculate ER2738 shaken in 25 ml of LB medium, , And the cells were incubated for 4 hours. Then, E. coli and phage were separated by centrifugation at 8,000 g. The phage-containing supernatant was treated with 6 ml of saline solution (20% PEG6000, 2.5 M NaCl) to precipitate the phage. After centrifugation at 8000G, the precipitated phages were treated with TBS solution to separate the phage.

Finally, the phage were amplified in a single colony to identify peptides with hair-binding phage. Escherichia coli ER2738 with phage was characterized by blue color in LB / X-gal / IPTG plate. To the ER2738 culture medium, 300 μl of the phage was diluted with TBS, treated with 2 μl, mixed with 4 ml of TOP agar, treated on the top of LB / X-gal / IPTG plate, and further cultured for 16 hours to select blue colonies. Each colony was further cultured in 5 ml of ER2738 culture medium for 6 hours, and DNA was isolated using M13 phage spin kit (Qiagen). Thus, peptides comprising the amino acid sequence of SEQ ID NO: 1 or 2 were selected.

SEQ ID NO: 1: KVWQLQLPQWIN

SEQ ID NO: 2: KVWTLSNDYIEF

Example  2: for biological structures Of peptide  Binding activity

The binding activity of the peptides to hair, skin and teeth was analyzed in order to confirm whether each peptide selected in Example 1 can exhibit binding activity to various bio-structures.

Example  2-1: Hair binding activity

Each of the peptides selected in Example 1 was chemically synthesized, and a peptide having the amino acid sequence of SEQ ID NO: 5 was used as a control. At this time, a Dansyl group was added to the C-terminal of each synthesized peptide.

SEQ ID NO: 5: TDMNKTEIRFVR

On the other hand, 50 mg of hair of a healthy adult was minced and immersed in a reaction buffer, and each of the synthesized peptides was added and then bound. The hair was then washed three times and neutralized by elution of the peptide in hair with 0.2 N HCl solution. The amount of peptides bound to the hair was measured with a victor fluorescence meter (340/520) and the binding activity of the peptides to the hair was analyzed (Table 1).

Amount of peptide bound to hair sample Fluorescence intensity (340/520) Control group
SEQ ID NO: 1
SEQ ID NO: 2 140
8200
4000

As shown in Table 1, the peptides of the control group were hardly bound to the hair, but the peptides of SEQ ID NO: 1 or 2 bound to the hair in a large amount, and in particular, the peptides of SEQ ID NO: I could.

Example  2-2: Skin binding activity

The binding activity of peptides to the skin was analyzed by the same method as in Example 2-1, except that pig skin was used instead of hair (Table 2).

The amount of peptide bound to the skin sample Fluorescence intensity (340/520) Control group
SEQ ID NO: 1
SEQ ID NO: 2 1500
72000
54000

As shown in Table 2, the peptide of the control group was hardly bound to the skin, but the peptide of SEQ ID NO: 1 or 2 bound to the skin in a large amount, and in particular, the peptide of SEQ ID NO: I could.

Example  2-3: tooth binding activity

The binding activity of peptides to teeth was analyzed by the same method as in Example 2-1 except that hydroxyapatite (HAP) powder, which is the core of the tooth structure, was used in place of the hair (Table 3) .

The amount of peptide bound to HAP sample Fluorescence intensity (340/520) Control group
SEQ ID NO: 1
SEQ ID NO: 2 1500
82000
40000

As shown in Table 3, the peptide of the control group was hardly bound to HAP, but the peptide of SEQ ID NO: 1 or 2 bound to HAP in a large amount, and in particular, the peptide of SEQ ID NO: 1 bound to HAP in the largest amount I could.

As shown in Examples 2-1 to 2-3, the peptides of the control group were hardly bonded to hair, skin and teeth. However, the peptides of SEQ ID NO: 1 or 2 provided in the present invention were found in large amounts in hair, skin and teeth And in particular peptides of SEQ ID NO: 1 bound to hair, skin and teeth in the greatest amount.

Therefore, it was found that the peptides of SEQ ID NO: 1 or 2 provided in the present invention exhibit binding activity to biological structures in general.

Example  3: For biological structures Of peptide As a crosslinking agent  activation

In order to confirm whether each of the peptides selected in Example 1 can exhibit activity as a crosslinking agent capable of binding a useful component to various biological structures, the activity of the peptide as a crosslinking agent in hair, skin and teeth was analyzed.

Example  3-1: For hair As a crosslinking agent  activation

Each of the peptides selected in Example 1 and the peptide having the amino acid sequence of SEQ ID NO: 5 was bound to the N-terminus of the GFP protein, and each GFP-fusion peptide in which the histag was bound to the C-terminus was prepared .

On the other hand, 50 mg of healthy adult hair was minced and immersed in a reaction buffer, and each of the GFP-fusion peptides prepared above was added and then bound. Then, the hair was washed three times, and reacted with anti-His antibody HRP (Horseradish peroxidase) conjugate for 5 minutes. Then, the hair was washed 10 times at 12,000 rpm for 1 minute to remove residual buffer. Then, the reaction was terminated by adding TMB (Tetramethyl benzidine) solution (Amersham), and the reaction was terminated by adding 0.2 N sulfuric acid. The absorbance (450 nm) was measured and the absorbance of the control group was subtracted from the absorbance By comparison, the binding activity of GFP-fusion peptide to hair was analyzed (Table 4).

The amount of GFP-fusion peptide bound to the hair sample Fluorescence intensity (450) Control group
SEQ ID NO: 1
SEQ ID NO: 2 0.015
0.982
0.753

As shown in Table 4, the GFP-fusion peptide of the control group was hardly bound to the hair, but the GFP-fusion peptide containing the peptide of SEQ ID NO: 1 or 2 was bound to the hair in a large amount. In particular, the peptide of SEQ ID NO: It was found that the contained GFP-fusion peptide binds to the hair in the largest amount.

Example  3-2: For skin As a crosslinking agent  activation

The binding activity of GFP-fused peptides to the skin was analyzed by the same method as in Example 3-1, except that pig skin was used instead of hair (Table 5).

The amount of GFP-fusion peptide bound to the skin sample Fluorescence intensity (450) Control group
SEQ ID NO: 1
SEQ ID NO: 2 0.012
1.082
0.853

As shown in Table 5, the GFP-fusion peptide of the control group was hardly bound to the skin, but the GFP-fusion peptide containing the peptide of SEQ ID NO: 1 or 2 was bound to the skin in a large amount and in particular, the peptide of SEQ ID NO: It was found that the GFP-fused peptides involved contained the greatest amount of binding to the skin.

Example  3-3: About teeth As a crosslinking agent  activation

The binding activity of GFP-fused peptides to teeth was analyzed by the same method as in Example 3-1, except that HAP powder, which is the core of the tooth structure, was used in place of the hair (Table 6).

The amount of GFP-fusion peptide bound to HAP sample Fluorescence intensity (450) Control group
SEQ ID NO: 1
SEQ ID NO: 2 0.011
1.282
0.743

As shown in Table 6, the GFP-fusion peptide of the control group was hardly bound to HAP, but the GFP-fusion peptide containing the peptide of SEQ ID NO: 1 or 2 was bound to HAP in a large amount and in particular, the peptide of SEQ ID NO: Containing GFP-fusion peptide bound to HAP in the highest amount.

As shown in Examples 3-1 to 3-3, the GFP-fusion peptide containing the peptide of the control group was hardly bound to hair, skin and teeth, but contained the peptide of SEQ ID NO: 1 or 2 provided in the present invention GFP-fusion peptide binds to hair, skin and teeth in a large amount, and in particular, GFP-fusion peptide including peptide of SEQ ID NO: 1 binds to hair, skin and teeth in the greatest amount.

Therefore, it was found that the peptide of SEQ ID NO: 1 or 2 provided in the present invention shows activity as a cross-linking agent capable of binding a useful substance to a biological structure in general.

<110> LG HOUSEHOLD & HEALTH CARE LTD. <120> Novel isolated peptide for binding to somatic structure and uses          the <130> PA130524 / KR <160> 5 <170> Kopatentin 2.0 <210> 1 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> peptide <400> 1 Lys Val Trp Gln Leu Gln Leu Pro Gln Trp Ile Asn   1 5 10 <210> 2 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> peptide <400> 2 Lys Val Trp Thr Leu Ser Asn Asp Tyr Ile Glu Phe   1 5 10 <210> 3 <211> 36 <212> DNA <213> Artificial Sequence <220> Coding sequence <400> 3 aaggtttggc agctgcagct gccgcagtgg attaat 36 <210> 4 <211> 37 <212> DNA <213> Artificial Sequence <220> Coding sequence <400> 4 taaggtgtgg acgctgtcga atgattatat tgagttt 37 <210> 5 <211> 12 <212> PRT <213> Artificial Sequence <220> <223> peptide <400> 5 Thr Asp Met Asn Lys Thr Glu Ile Arg Phe Val Arg   1 5 10

Claims (11)

A novel isolated peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, exhibiting bio-structure binding activity.
The method according to claim 1,
Wherein the bio-structure is hair, skin or teeth.
3. A polynucleotide encoding the peptide of claim 1 or 2.
The method of claim 3,
Wherein the polynucleotide comprises the nucleotide sequence of SEQ ID NO: 3 or SEQ ID NO: 4.
An expression vector comprising the polynucleotide of claim 3.
A transformant wherein the expression vector of claim 5 is introduced into a host cell.
(a) obtaining a polynucleotide encoding the amino acid sequence of SEQ ID NO: 1 or 2;
(b) cloning the obtained polynucleotide to obtain an expression vector;
(c) introducing the obtained expression vector into a host cell to obtain a transformant; And
(d) culturing the transformant and recovering the desired peptide therefrom. &lt; Desc / Clms Page number 20 &gt;
8. The method of claim 7,
Wherein the polynucleotide comprises the nucleotide sequence of SEQ ID NO: 3 or 4.
A crosslinking agent for hair, skin or teeth comprising the peptide of claim 1 or 2.
10. A composition for delivery to hair, skin or teeth, comprising the crosslinker of claim 9.
11. The method of claim 10,
Wherein the composition is selected from the group consisting of pharmaceutical compositions, quasi-drug compositions and cosmetic compositions.