CN1986562A - Polypeptide segment capable of enhancing animal's antibody response and its application - Google Patents
Polypeptide segment capable of enhancing animal's antibody response and its application Download PDFInfo
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- CN1986562A CN1986562A CN 200610147255 CN200610147255A CN1986562A CN 1986562 A CN1986562 A CN 1986562A CN 200610147255 CN200610147255 CN 200610147255 CN 200610147255 A CN200610147255 A CN 200610147255A CN 1986562 A CN1986562 A CN 1986562A
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- mouth disease
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Abstract
The present invention belongs to the field of immunology and genetic engineering technology, and is especially one polypeptide segment capable of enhancing animal's antibody response level and its application. The polypeptide segment can stimulate genetic engineering vaccine to induce response level of neutralizing antibody, and is especially the 1-33 positions amino acid sequence of foot-and-mouth disease virus VP2 protein. The present invention adopts beta-galactosidase as carrier protein and connects the said immune stimulating polypeptide segment to the C end of the carrier protein to prepare fusion protein LacZ-B1. The fusion protein LacZ-B1 may be used as the immune stimulating adjuvant of foot-and-mouth disease genetic engineering polypeptide vaccine, and has high safety and capacity of raising animal's antibody response level.
Description
Technical field
The invention belongs to immunology and gene engineering field, be specifically related to a kind of polypeptide fragment and application thereof that can strengthen animal's antibody response.
Background technology
World Organization for Animal Health classifies foot and mouth disease first of the category-A transmissible disease as.Foot and mouth disease is cloven-hoofed animals such as livestock contagious disease the most serious in the world today, main harm pig, ox, sheep.For many years, foot and mouth disease is large-scale outbreak and popular worldwide, causes the tremendous economic loss to livestock industry.Preventive vaccination is the main means of this virus of control.In the vaccine of various prevention foot and mouth disease, recombinant vaccine is easy to preserve because security is good, and advantages such as effect stability have broad application prospects.A recombinant vaccine key of success is that can it cause that animal produces sufficiently high neutralizing antibody and replys.Proved that now foot and mouth disease virus VP1 Protein G-H ring is an important antigen site of inducing neutralizing antibody, related to 130-160 position peptide section.Yet the fusion rotein that contains this peptide section but can not bring out animal and produce sufficiently high neutralizing antibody and reply.Therefore, seeking a kind of this peptide section that can strengthen brings out the novel immunity enhancement adjuvant that animal produces the neutralizing antibody ability and seems particularly necessary.
Summary of the invention
The objective of the invention is to propose a kind of polypeptide fragment and application thereof that can strengthen animal's antibody response.
The polypeptide fragment that can strengthen animal's antibody response that the present invention proposes is a kind ofly can bring out the immunostimulation polypeptide fragment that neutralizing antibody is replied level by the stimulated gene engineered vaccine.This immunostimulation fragment is a foot and mouth disease virus VP2 albumen 1-33 amino acids, its aminoacid sequence is: DKKTEEATLLEDRILTTRNGHTTSTTQSSVGVT (SEQ ID NO.1), this peptide called after B1, veriform foot and mouth disease virus, this immunostimulation fragment has 1-3 amino acid to be replaced.
The immunostimulation fragment that the present invention proposes confirms that with western blotting and T cell proliferation experiment this fragment contains the B cell epitope, has T cytositimulation function.This segmental immune-stimulating effect is that mainly it strengthens t cell response in the animal body, further promotes the B cell maturation in differentiation, has strengthened the secretion of antibody.
The present invention also adopts beta-galactosidase enzymes as carrier proteins, above-mentioned immunostimulatory peptides section is connected this carrier proteins tilactase C end, be prepared into fusion rotein LacZ-B1, this fusion rotein can be used as novel foot-and-mouth disease gene engineering polypeptide vaccine immunostimulation adjuvant.This fusion rotein adjuvant security is good, can effectively improve the animal's antibody response level.
Description of drawings
Fig. 1: expressing fusion protein and immunogenicity thereof detect.
The A fusion rotein is expressed SDS-PAGE the JM101 bacterium
M: molecular weight of albumen Marker.
The empty bacterium contrast of 1:JM101 2:pWR590 contrast 3:pWRB1
B Western blotting detects fusion protein immunization originality
1-3 is same as described above
Fig. 2 adopts the MTS method to measure T cell proliferation.
Embodiment
Embodiment 1, is that carrier proteins makes up immunostimulation albumen pWRB1 expression plasmid with the beta-galactosidase enzymes.Its result is as follows:
SDS-PAGE electrophoresis detection proof is expressed the fusion rotein LacZ-B1 molecular weight and expection consistent (seeing Figure 1A) that obtains.Western blotting detects this LacZ-B1 fusion rotein of proof and has very strong antigenicity, can special antigen antibody reaction (seeing Fig. 1 (B)) take place with standard antibody.
With this fusion protein immunization cavy, detect T cell proliferation.The result proves among the VP2 1-33 peptide section induced animal T cell proliferation (see figure 2) effectively.X-coordinate among the figure is represented standard antigen extension rate X, ordinate zou is represented stimulation index SI (annotate: this experiment adopts the MTS/PMS method to measure T cell proliferation, stimulation index is with the OD value of experimental group and do not add the difference of antigenic negative control group OD and do not add the ratio of antigenic negative control group OD value).
Fusion rotein LacZ-B1 preparation vaccine with purifying, (Asia I type foot and mouth disease virus VP1 albumen 133-163 peptide section is connected in beta-galactosidase enzymes C and holds formed fusion rotein this vaccine and LacZ-D7 vaccine, this fusion protein immunization cavy, can cause that the cavy neutralizing antibody is replied sees Table 1) twice immune guinea pig (immunizing dose: each component protein single immunization 300 μ g) of associating, compare with independent immune LacZ-D7 fusion rotein group, the former significantly improves (seeing Table 2) than the latter at caused neutralizing antibody level.
Table 1 fusion rotein immune guinea pig neutralizing antibody is respectively measured
| Group number | NAT (PD 50) | |||||
| 1 | 2 | 3 | 4 | 5 | 6 | |
| Control LacZ LacZ-D7 LacZ-B1 | 0 0 91 0 | 0 0 256 0 | 0 0 256 0 | 0 0 64 0 | 0 0 90 0 | 0 0 76 0 |
Table 2 fusion rotein mixes the immune guinea pig neutralizing antibody and measures
| Group number | Each cavy NAT | P value | |||||||||||
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 | ||
| D7 (LacZ-D7)+LacZ (LacZ-D7)+(LacZ-B1) | 65 362 724 | 90 362 724 | 76 180 1448 | 215 362 256 | 32 215 512 | 256 64 1024 | 362 0 1024 | 48 - 235 | 1024 - 1024 | 91 - 96 | 256 - 1024 | 256 - 1024 | AC 0.416093 0.001484 |
Sequence table:
SEQID NO.1：DKKTEEATLLEDRILTTRNGHTTSTTQSSVGVT
Claims (3)
1. polypeptide fragments that can strengthen animal's antibody response, it is characterized by this polypeptide fragment and be connected in carrier proteins tilactase C end, constitute a fusion rotein, described polypeptide fragment is a foot and mouth disease virus VP2 albumen 1-33 amino acids, and its aminoacid sequence is: DKKTEEATLLEDRILTTRNGHTTSTTQSSVGVT.
2, polypeptide fragments according to claim 1 is characterized in that this polypeptide fragments has 1-3 amino acid to be replaced.
3, the application of a kind of polypeptide fragments as claimed in claim 1 or 2 in the immunostimulation adjuvant of preparation foot-and-mouth disease gene engineering polypeptide vaccine, this adjuvant are as carrier proteins, be connected this carrier proteins tilactase C end and the fusion rotein that obtains by described polypeptide fragments with beta-galactosidase enzymes.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN200610147255A CN1986562B (en) | 2006-12-14 | 2006-12-14 | Polypeptide segment capable of enhancing animal's antibody response and its application |
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| CN200610147255A CN1986562B (en) | 2006-12-14 | 2006-12-14 | Polypeptide segment capable of enhancing animal's antibody response and its application |
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| Publication Number | Publication Date |
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| CN1986562A true CN1986562A (en) | 2007-06-27 |
| CN1986562B CN1986562B (en) | 2010-05-19 |
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| CN200610147255A Expired - Fee Related CN1986562B (en) | 2006-12-14 | 2006-12-14 | Polypeptide segment capable of enhancing animal's antibody response and its application |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103255159A (en) * | 2013-03-04 | 2013-08-21 | 张掖市奥林贝尔生物科技有限公司 | Recombinant expression and preparation method of cecropin CAD |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1470285A (en) * | 2003-06-13 | 2004-01-28 | 复旦大学 | Polypeptide vaccine of anti Asiatic I virus of foot-and-mouth disease and its preparing method |
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2006
- 2006-12-14 CN CN200610147255A patent/CN1986562B/en not_active Expired - Fee Related
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103255159A (en) * | 2013-03-04 | 2013-08-21 | 张掖市奥林贝尔生物科技有限公司 | Recombinant expression and preparation method of cecropin CAD |
| CN103255159B (en) * | 2013-03-04 | 2015-11-18 | 张掖市奥林贝尔生物科技有限公司 | A kind of cecropin CAD's is recombinant expressed and preparation method thereof |
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| CN1986562B (en) | 2010-05-19 |
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