CN102060912B - Amyloid protein oligomer conformation type epitope polypeptide and application thereof - Google Patents
Amyloid protein oligomer conformation type epitope polypeptide and application thereof Download PDFInfo
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Abstract
The invention belongs to the field of peptide-containing medical preparations, in particular relating to an amyloid protein oligomer conformation type epitope polypeptide and an application thereof. The epitope polypeptide is used for preparing an antibody which can identify a plurality of amyloid protein oligomers for the diagnosis, treatment and experimental research of amyloid protein diseases. The invention provides the amyloid protein oligomer conformation type epitope polypeptide (PEOZ) the amino acid sequence of which is Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys or is obtained by replacing, deleting or adding one or more of amino acid residues to the amino acid sequence. The epitope polypeptide can effectively stimulate an organism to generate the specific antibody which can identify amyloid protein oligomers and the antibody can inhibit the aggregation of the amyloid protein and the cytotoxicity.
Description
Technical field
The invention belongs to the pharmaceutical product field that contains peptide, be specifically related to a kind of amyloid oligomer conformation type antigen epitope polypeptide and application thereof; The present invention is applied to discern diagnosis, treatment and the experimental study of the Antibody Preparation amyloid disease of multiple amyloid oligomer.
Background technology
For many years, the multiple disease that causes of the gathering of amyloid or polypeptide self has caused increasing harm to human beings'health.Nontoxicity of some protein own or toxicity are very little; But they self can be gathered into oligomer (oligomer) with toxic action or fibrous material (fibril) and cause a series of diseases; As the A Zihaimo that causes by Beta-amyloid (A-Beta) sick (being commonly called as senile dementia) (Alzheimer ' disease; AD); The Parkinson's disease that cause by alpha-synuclein (Parkinson ' disease, PD), at least ten surplus kinds of encephalopathics of the humans and animals that comprises mad cow disease that causes by prion protein (Prion protein (PrP)); At least 9 kinds of heredity nerve degenerative diseases that comprise Huntington Chorea (Huntington ' s disease) that cause by the polypeptide that contains poly glumine (PolyQ); (islet amyloid polypeptide, IAPP, type ii diabetes that amylin) causes and the N,O-Diacetylmuramidase (lysozyme) that is caused by long-time dialysis assemble the disease that deposition causes etc. by islet amyloid polypeptide.Wherein, to human health risk maximum be AD and PD and type ii diabetes.Medical statistics shows, 5～6% suffer from AD among the over-65s the elderly in China and the American-European countries, and sickness rate rises year by year.This disease has been classified as causes dead the fourth-largest disease, is only second to heart trouble, cancer and apoplexy.Other have an appointment 1% over-65s the elderly suffers from PD.And the number of suffering from type ii diabetes can reach more than 5% of total population.These diseases have caused increasing harm to human beings'health, and the basic reason of its morbidity (or partly cause) is the gathering of amyloid or polypeptide self.
Research shows that the gathering of different proteins starts from protein (or polypeptide) monomer of false folding or sex change at first, and the formation of monomer polypeptide interchain hydrogen bond has caused the polymerization of protein molecular.At first form spherical oligomer by the solubility of electronics or the about 3-10 nm of the observable size of AFM; Some oligomer can further be combined into crooked flexible silk (protofibril), and then the formation diameter is the smooth surface of 6-10 nm or fiber in the shape of a spiral.The protein of non-homology finally can form the protein polymer with analog structure.The fiber that is formed by various amyloid aggregations such as A-Beta, alpha-synuclein and amylin all contains " cross-Beta " structure, and wherein the skeleton of Beta lamella formation is vertical with the fiber longitudinal axis, and the hydrogen bond net in the skeleton is parallel with the longitudinal axis.Polypeptide is arranged as parallel Beta chain in the Beta lamella, in the Beta lamella, amino acid all has accurate position.Even the oligomer of different sources also has similar constitutional features, the oligomer that oligomer specific recognition antibody can form with the amyloid monomer by different sources combines, and does not combine with their monomer and fiber.This shows that oligomer can form the distinctive epitope of common oligomer that is independent of outside its amino acid primary sequence by the protein polypeptide skeleton.Colloid gold particle simulation A-Beta 40 oligomers that the Glabe laboratory applications is connected with A-Beta 40 prepared not only can combine with A-Beta 42 oligomers with A-Beta 40 and can with the polyclonal antibody (A11) of the oligomer specific combination of formation such as alpha-synuclein, IAPP, PolyQ, PrP, Insulin, Lysozyme, this antibody can also effectively suppress the cytotoxicity of all these oligomers.
What the past people thought the amyloid disease always is to be caused by the insoluble fiber that protein aggregation becomes.A large amount of in recent years researchs show that the crucial paathogenic factor of many amyloid diseases is the less water-soluble oligomeric things of volume.The disease that is caused by the albumen oligomer exists similar mechanism, and promptly cell membrane damage, oxidative stress, mitochondrial function imbalance, necrocytosis, signal transmission are unusual etc.But people it be unclear that the detailed mechanism that oligomer influences the normal physiological activity of cell.How are the topological framework of oligomer and epitope thereof, and these all are that people demand the problem inquiring into and solve urgently how could to suppress its cytotoxicity etc. effectively.Thereby use the gathering that active and passive immunological technique treatment AD suppress A-Beta, and the removing of quickening A-Beta is one of focus of AD research field.
2000-2002 U.S. scientific research personnel uses the vaccine that A-Beta processes and has carried out an animal experiment and a clinical trial phase.Experimental result is encouraging, and laboratory animal and patient's clinical symptom obviously alleviates, and memory is obviously recovered, and pathological change is also significantly gone down.Simultaneously, use anti-A-Beta antibody the passive immunotherapy of laboratory animal has also been received better curative effect.Yet, unfortunately in phase ii clinical trial,, have 6% patient table to reveal meningitic symptom and pathological change, the patient who has even take place dead though most patient still shows good result.Therefore, the experimental study to A-Beta vaccine and antibody stops immediately.However, we can find out that the A-Beta vaccine should affirm the result of treatment of AD from above-mentioned research.Research shows that the spinoff that is caused by the A-Beta vaccine possibly be the autoimmune response by the T cell mediated of this vaccine-induced body generation, and the generation of this spinoff is caused by Th1 rather than the immunoreation of Th2 type.In order to overcome the spinoff of A-Beta vaccine, when vaccine design, should strengthen bringing out B cell response and the immunoreactive generation of Th2 type, and avoid bringing out t cell immune response and the immunoreactive generation of Th1 type.Vaccine based on the design of A-Beta oligomer epitope peptide can easily be taken into account above-mentioned characteristic, also can not induce body to form the monomeric antibody of anti-A-Beta.Back one characteristic can thoroughly be eliminated the possibility that has side effects between the A-Beta monomer in vaccine and the body, and can not influence the normal physiological function of A-beta.Therefore, the peptide vaccine of developing according to A-Beta oligomer epitope will have the potential using value, and obtain the prerequisite that A-Beta oligomer epitope is this peptide vaccine design and development.So, scientific research and clinical in all need obtain a kind of amyloid oligomer conformation type antigen epitope polypeptide.
Summary of the invention
The object of the present invention is to provide a kind of amyloid oligomer conformation type antigen epitope polypeptide (being called for short PEOZ); Its aminoacid sequence is: Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys; Or by above-mentioned aminoacid sequence through replacement, disappearance or add one or several amino acid, and have the polypeptide that suppresses the amyloid aggregation effect.The present invention can produce the specific antibody of discerning the amyloid oligomer by the effective stimulus body, and this antibody can suppress the gathering and the cytotoxicity of amyloid.
In specific embodiment of the present invention; The N of polypeptide according to the invention end Using P EG2000 is modified (sequence is: PEG-Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys), the deletion L-Ala (sequence is: Cys-Ser-His-Leu-His-Asn-Asn-Cys), Serine be replaced as L-Ala (sequence is: Cys-Ala-His-Ala-Leu-His-Asn-Asn-Cys) or insert L-Ala (sequence is: Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys), through detect show the PEOZ polypeptide through some modification, certain amino acid is through displacement, disappearance or in polypeptide, still can remain with and A11 bonded ability behind the increase amino acid.
In vitro tests shows that polypeptide of the present invention can combine with A11, has proved that described polypeptide is one of pairing epitope of polyclone A11; Described polypeptide has good antigenic characteristic, but the effective stimulus body produces antibody; Described polypeptide is a common epitope on the multiple amyloid oligomer; Described polypeptide antibody can obviously suppress amyloid A b42, PrP, amylin, a-synuclein, lysozyme gathering; Described polypeptide antibody can obviously suppress the cytotoxicity of Ab42, PrP, amylin, a-synuclein, lysozyme.
The present invention also provides said polypeptide to be applied to amyloid oligomer suppressor factor.
The present invention also provides said polypeptide to be applied to prepare specific recognition amyloid oligomer antibody.
The present invention also provides said polypeptide to be applied to preparation treatment amyloid disease vaccine.
The present invention compares prior art and has following beneficial effect:
1. the present invention can produce the specific antibody of discerning the amyloid oligomer by the effective stimulus body, and this antibody can suppress the gathering and the cytotoxicity of amyloid.
2. B cell response and the immunoreactive generation of Th2 type be can strengthen bringing out based on vaccine of the present invention, and t cell immune response and the immunoreactive generation of Th1 type avoided bringing out; Can not induce body to form the monomeric antibody of anti-A-Beta, can thoroughly eliminate the possibility that has side effects between the A-Beta monomer in vaccine and the body, and can not influence the normal physiological function of A-beta.
3. the present invention has better immunogenicity, but the effective stimulus body produces antibody, thereby aspect the development of vaccine, can be used as good immunogen in amyloid treatment of diseases or prevention.
Description of drawings:
Fig. 1. PEOZ and A11 bonded ELISA measure;
Fig. 2. measure through replacing, lack or inserted amino acid whose PEOZ and A11 bonded ELISA;
Fig. 3. the antibody after the PEOZ immunity in the mice serum combines with the Ab42 oligomer;
Fig. 4. PEOZ antibody combines with the oligomer of multiple amyloid;
Fig. 5. ThT fluorescent dyeing determination PEOZ antibody suppresses the gathering of amyloid;
Fig. 6. PEOZ antibody suppresses the cytotoxicity of amyloid;
Embodiment:
Embodiment 1:
One peptide species (being called for short PEOZ); Its aminoacid sequence is: Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys; Or by above-mentioned aminoacid sequence through replacement, disappearance or add one or several amino acid, and have the polypeptide that suppresses the amyloid aggregation effect.The present invention can produce the specific antibody of discerning the amyloid oligomer by the effective stimulus body, and this antibody can suppress the gathering and the cytotoxicity of amyloid.
The N of said polypeptide end Using P EG2000 is modified (sequence is: PEG-Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys), the deletion L-Ala (sequence is: Cys-Ser-His-Leu-His-Asn-Asn-Cys), Serine be replaced as L-Ala (sequence is: Cys-Ala-His-Ala-Leu-His-Asn-Asn-Cys) or insert L-Ala (sequence is: Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys), through detect show the PEOZ polypeptide through some modification, certain amino acid is through displacement, disappearance or in polypeptide, still can remain with and A11 bonded ability behind the increase amino acid.
In vitro tests shows that described polypeptide can combine with A11, has proved that described polypeptide is one of pairing epitope of polyclone A11; Described polypeptide has good antigenic characteristic, but the effective stimulus body produces antibody; Described polypeptide is a common epitope on the multiple amyloid oligomer; Described polypeptide antibody can obviously suppress amyloid A b42, PrP, amylin, a-synuclein, lysozyme gathering; Described polypeptide antibody can obviously suppress the cytotoxicity of Ab42, PrP, amylin, a-synuclein, lysozyme.
Said polypeptide is applied to the amyloid polygalacturonase inhibitor; Also be applied to prepare specific recognition amyloid oligomer antibody; Also be applied to preparation treatment amyloid disease vaccine.
Said polypeptide can produce the specific antibody of discerning the amyloid oligomer by the effective stimulus body, and this antibody can suppress the gathering and the cytotoxicity of amyloid.B cell response and the immunoreactive generation of Th2 type be can strengthen bringing out based on the vaccine of said polypeptide, and t cell immune response and the immunoreactive generation of Th1 type avoided bringing out; Can not induce body to form the monomeric antibody of anti-A-Beta, can thoroughly eliminate the possibility that has side effects between the A-Beta monomer in vaccine and the body, and can not influence the normal physiological function of A-beta.
The method of the synthetic preparation of PEOZ is:
PEOZ is annular seven peptides; (preparation method is referring to Weng C C by the synthetic preparation of ordinary method by the biochemical ltd of Shanghai gill; Peter D W. Fmoc Solid Phase Peptide Synthesis:A Practical Approach. Oxford:University Press, 2000. 1 ~ 8; Atherton E, Sheppard R C. Solid Phase Peptide Synthesis:A Practical Approach. Oxford:IRL Press, 1989); PEOZ purity is more than or equal to 95%; PEOZ is stored in-20 ° of C, avoids multigelation.
The preparation method that PEOZ is used for special anti-amyloid oligomer antibody is:
With the M13 phage immunity new zealand white rabbit of showing the PEOZ polypeptide.Per 2 all immunity once, it be an individual phagemid/rabbit that antigen is used in each immunity.Immunity is 4 times altogether.After last immune 15 days, gather rabbit anteserum.Sepharose to be connected with the PEOZ polypeptide carries out affinity chromatography to rabbit anteserum, the antibody of the anti-PEOZ of purifying.The antibody of the oligomer of this specific combination amyloid can be used for experimental study, and amyloid treatment of diseases and clinical diagnosis are also had the potential using value.
All experimental datas of PEOZ are all through at least 3 independently experiment acquisitions, and experimental data is expressed as mean number plus-minus standard deviation.The statistical study of data is to use One-way ANOVA software to carry out.
Below in conjunction with embodiment 2-7, further set forth the character of PEOZ.
Embodiment 2:PEOZ combines with polyclonal antibody A11
100 μ l PEOZ (1 mg/ hole) are encapsulated 96 hole enzyme linked immunological microwell plates (with the hole that do not encapsulate PEOZ as contrast), put 37 ° of C 2 h, seal 2 h with 37 ° of C of 3% BSA, 200 μ L/ holes.Wash plate 3 times with PBS.Every hole adds 100 μ l A11 (1:500), behind incubated at room 1 h, with the PBS washing that contains 0.1% Tween-20 3 times.Behind incubated at room 1 h, with the PBS washing that contains 0.1% Tween-20 3 times.Every hole adds the goat-anti rabbit HRP ELIAS secondary antibody (1:5000) after the 100 μ l dilution, behind incubated at room 1 h, with the PBS washing that contains 0.1% Tween-20 3 times.Every hole adds substrate solution TMB 100 μ L, places 37 ° of C 20 min, and every then hole adds 50 μ l, 1 mmol/L sulfuric acid termination reaction, on enzyme-linked immunosorbent assay instrument, measures OD value (wavelength 450nm).PEOZ shown in Figure 1 can be by the A11 specific recognition.PEOZ is the polypeptide that obtains as the screening substrate with A11, it with A11 combine confirmed that it is one of pairing epitope of polyclonal antibody A11.
Embodiment 3: through displacement, lack or inserted amino acid whose PEOZ and still combine with A11
The N of PEOZ end Using P EG2000 is modified (abbreviating PEG-PEOZ as), and (sequence is: PEG-Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys); (abbreviate as: PEOZ-A) (sequence is the deletion L-Ala: Cys-Ser-His-Leu-His-Asn-Asn-Cys); Serine is replaced as L-Ala (abbreviating PEOZ-S/A as), and (sequence is: Cys-Ala-His-Ala-Leu-His-Asn-Asn-Cys) or insert L-Ala (abbreviate as: PEOZ-G) (sequence is: Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys).In order to detect the situation that combines of improved polypeptide and A11, with these polypeptide with have the histidine-tagged PEOZ that does not transform and combine to competition with A11, then with the combine situation of the ELISA detection latter with A11.Add and be coated with in the hole of A11 adding the PEOZ that has label that 1 mg do not transform mixture 100 ml, detect with ELISA then and have the PEOZ of label and combining of A11 with the improved PEOZ of 10 mg, mensuration OD value, the result sees Fig. 2.Show that the PEOZ polypeptide still can combine A11 with the PEOZ effective competition that unmodified is transformed through some modification, certain amino acid through displacement, disappearance or after in polypeptide, increasing amino acid; Cause the PEOZ that has label obviously to reduce (compare, P < 0.01) with the combination of A11 with independent PEOZ positive control.
Embodiment 4:PEOZ can bring out body and produce antibody
Phage (as the contrast) immune mouse (5/group) of using the M13 phage of showing the PEOZ polypeptide and not having polypeptide to show, every each immune phage 10
10Individual.Per 2 all immunity once are total to immunity 4 times.The 4th immunity blood sampling in back 15 days, separation of serum.Use the content that ELISA detects PEOZ antibody in the serum.100 μ l Ab oligomer (1 mg/ hole) are encapsulated 96 hole enzyme linked immunological microwell plates, put 37 ° of C 2 h, seal 2 h (200 μ L/ hole) with 37 ° of C of 3% BSA.Wash plate 3 times with PBS, add the serum of 2000 times of dilutions, behind incubated at room 1 h, with the PBS washing that contains 0.1% Tween-20 3 times.Every hole adds the sheep anti mouse ELIAS secondary antibody (1:3000) after the 100 μ l dilution, behind incubated at room 1 h, with the PBS washing that contains 0.1% Tween-20 3 times.Every hole adds substrate solution TMB 100 μ L, places 37 ° of C 20min, and every then hole adds 50 μ l, 1 mmol/L sulfuric acid termination reaction, on enzyme-linked immunosorbent assay instrument, measures OD value (wavelength 450nm) (Fig. 3).5 mouse of immune PEOZ shown in Figure 2 can both produce the antibody of anti-PEOZ, show that PEOZ has good antigenic characteristic, but the effective stimulus body produces antibody.
The preparation of the immune antibody of embodiment 5:PEOZ and with the combining of each amyloid oligomer
With the M13 phage immunity new zealand white rabbit of showing the PEOZ polypeptide.Per 2 all immunity once, each antigen 10
12Individual phagemid.Immunity is 4 times altogether.After last immune 15 days, gather rabbit anteserum.Sepharose to be connected with the PEOZ polypeptide carries out affinity chromatography to rabbit anteserum, the antibody of the anti-PEOZ of purifying.
100 μ l Ab42, PrP, amylin, a-synuclein, lysozyme monomer (mon), oligomer (olig) and fiber (fib) (1 mg/ hole) are encapsulated 96 hole enzyme linked immunological microwell plates, the negative contrast of BSA respectively.Anti-with PEOZ antibody as one, anti-with the goat-anti rabbit LgG of HRP mark as two, and by the operation that experimentizes of above-mentioned ELISA method.Fig. 4 shows; PEOZ antibody can combine with the oligomer of Ab, PrP, amylin, a-synuclein respectively; And do not combine (except the lysozyme) with their monomer and fiber; Show that PEOZ is a common epitope on the multiple amyloid oligomer, and the monomer of amyloid and fiber there is not this epitope (except the lysozyme) (Fig. 4).
The gathering of embodiment 6:PEOZ antibody vitro inhibition amyloid
1) Ab42, amylin, the a-synuclein with foreign procurement is dissolved to 1 mg/ml with hexafluoroisopropanol (HFIP), and room temperature ultrasonic is handled 10 min, divides to install in the epidorf pipe, with the HFIP that volatilizees in the vacuum, preserves in-20 ° of C then.Place 20 min with preceding Ab42, amylin, the a-synuclein that HFIP was handled in room temperature, add DMSO 99.8MIN. (DMSO) then, making each protein concentration is 5 mg/ml, and the PBS damping fluid with 0.02 M pH 7.4 is diluted to desired concn then.PrP, lysozyme then directly are prepared into desired concn with 0.02 M pH, 7.4 PBS.
2) PEOZ antibody is dissolved in the PBS damping fluid of 0.02 M pH 7.4; Join Ab42, PrP, amylin, a-synuclein then, (lysozym is the acetate buffer solution of pH2.5 to lysozym solution; All the other samples are the PBS damping fluid of pH7.4) in, make each proteic final concentration be respectively 10,200,20,40,200 μ M.The mol ratio of each albumen and PEOZ antibody is respectively 1:1,10:1,1:1,2:1 and 10:1.With the protein solution that do not add PEOZ antibody as contrast.Ab42, PrP, amylin, a-synuclein, lysozym sample were placed respectively 1 day, 5 days, 5 hours, 4 days and 6 days in 37 ° of C (lysozym is positioned over 65 ° of C).
3) it is 5 mM that the phosphate buffered saline buffer that thioflavine (ThT) is dissolved in pH 6.5,50 mM makes its concentration.Sample 20 ml that get after hatching join in the black enzyme plate that contains 180 ml ThT solution.On multi-functional ELIASA, measure the fluorescence intensity of ThT behind the mixing with the emission wavelength of 450 nm excitation wavelengths and 482 nm.The fluorescence intensity of each sample is deducted the background fluorescence of ThT itself and the significant difference situation between the analytic sample.Add the albumen fluorescence intensity that reduces behind the PEOZ antibody and be the inhibiting rate of antibody protein aggregation divided by the albumen fluorescence intensity that does not add antibody.Fig. 5 is the gathering inhibiting rate of PEOZ antibody to Ab42, amylin, a-synuclein, lysozyme, PrP, shows that PEOZ can obviously suppress the gathering of each amyloid.
Embodiment 7:PEOZ antibody suppresses the cytotoxicity of amyloid
Substratum (MEM) with containing 10% foetal calf serum is made into the individual cells suspension with the SH-SY5Y cell, with 10000 cell inoculations in every hole to 96 porocyte culture plates, every pore volume 100 μ L.Cell was cultivated 24 hours in 37 ° of C, and incubator CO2 concentration is 5%.Every hole adds the sample (mixture of Ab42, PrP, amylin, a-synuclein, lysozyme and PEOZ antibody; Amyloid contrast and PBS contrast); Ab42, PrP, amylin, a-synuclein, the proteic final concentration of lysozyme are to be respectively 2,200,5,2,20 μ M, and the final concentration of PEOZ antibody is 1,20,2.5,1,10 μ M.Cell continued to cultivate after 48 hours, and every hole adds MTT solution (5 mg/mL) 10 μ L, and 37 ° of C were hatched 3 hours; Stop cultivating; Every hole adds 100 μ L dissolution of crystals liquid (10%SDS and 5% isopropylcarbinol are dissolved among the 0.01 M HCL), and 37 ° of C incubated overnight are fully dissolved crystallisate.On multi-functional enzyme-linked immunosorbent assay instrument, measure each hole absorbance value with the 570nm wavelength.With the absorbancy of sample divided by the absorbancy of the cell that does not add sample as the cell activity index, and the significant difference situation between the analytic sample (compare with each amyloid contrast,, P < 0.05; #, P 0.01), the result shows that PEOZ antibody can obviously suppress the cytotoxicity (Fig. 6) of Ab42, PrP, amylin, a-synuclein, lysozyme.
The above only is a preferred implementation of the present invention; Should be pointed out that for those skilled in the art, under the prerequisite that does not break away from the principle of the invention; Can also make some improvement and retouching, these improvement and retouching also should be regarded as protection scope of the present invention.
< 110>Tsing-Hua University
<120>
<160>?5
<210>?1
<211>?9
<212>?PRT
< 213>artificial sequence
<400>?1
Cys?Ser?His?Ala?Leu?His?Asn?Asn?Cys
<210>?2
<211>?9
<212>?PRT
< 213>artificial sequence
<220>
< 223>aminoterminal is modified with PEG2000
<400>?2
Cys?Ser?His?Ala?Leu?His?Asn?Asn?Cys
<210>?3
<211>?8
<212>?PRT
< 213>artificial sequence
<400>?3
Cys?Ser?His?Leu?His?Asn?Asn?Cys
<210>?4
<211>?9
<212>?PRT
< 213>artificial sequence
<400>?4
Cys?Ala?His?Ala?Leu?His?Asn?Asn?Cys
<210>?5
<211>?9
<212>?PRT
< 213>artificial sequence
<400>?5
Cys?Ser?His?Ala?Leu?His?Asn?Asn?Cys
Claims (8)
1. an amyloid oligomer conformation type antigen epitope polypeptide is characterized in that its aminoacid sequence is: Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys.
2. polypeptide according to claim 1 is characterized in that, described polypeptide N end is modified with PEG2000.
3. an amyloid oligomer conformation type antigen epitope polypeptide is characterized in that its aminoacid sequence is: Cys-Ser-His-Leu-His-Asn-Asn-Cys.
4. an amyloid oligomer conformation type antigen epitope polypeptide is characterized in that its aminoacid sequence is: Cys-Ala-His-Ala-Leu-His-Asn-Asn-Cys.
5. an amyloid oligomer conformation type antigen epitope polypeptide is characterized in that its aminoacid sequence is: Cys-Ser-His-Ala-Leu-His-Asn-Asn-Cys.
6. be applied to prepare amyloid oligomer suppressor factor according to each said polypeptide of claim 1-5.
7. be applied to prepare specific recognition amyloid oligomer antibody according to each said polypeptide of claim 1-5.
8. be applied to preparation treatment amyloid disease vaccine according to each said polypeptide of claim 1-5.
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| CN101736004A (en) * | 2008-11-18 | 2010-06-16 | 复旦大学 | Method for preparing high-purity amyloid polypeptide Abeta (1-40) |
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