70 results about "Cholera Toxin B Subunit" patented technology

This invention relates in part to synthesizing high value pharmaceutical proteins in transgenic plants by chloroplast expression for pharmaceutical protein production. We use poly(GVGVP), for example, as a fusion protein to enable hyper-expression of insulin and to accomplish rapid one step purification of fusion peptides utilizing the inverse temperature transition properties of this polymer. We also use insulin-CTB fusion protein in chloroplasts of nicotine free edible tobacco (LAMD 605) for oral delivery. This invention includes expression of native cholera toxin B subunit gene as oligomers in transgenic tobacco chloroplasts which may be utilized in connection with large-scale production of purified CTB, as well as an edible vaccine if expressed in an edible plant, as a transmucosal carrier of peptides to which it is fused to enhance mucosal immunity, and/or to induce oral tolerance of the products of these peptides. The present invention also relates in part to recombinant DNA vectors for enhanced expression of human serum albumin, insulin-like growth factor I, and interferon-α 2 and 5, via chloroplast genomes.

The invention provides a helicobacter pylori multiepitope vaccine. The active ingredient of the helicobacter pylori multiepitope vaccine is a piece of polypeptide; and the helicobacter pylori multiepitope vaccine mainly comprises a multi-copy body of Th and B cell antigen epitopes of helicobacter pylori urease A and B bi-subunit and a mucosal immune adjuvant of a cholera toxin B subunit. The preparation method mainly comprises the following steps of: synthesizing an artificial gene by using a gene synthesis technology, wherein the artificial gene comprises a gene sequence of the multi-copy body of the Th and B cell antigen epitopes of the helicobacter pylori urease A and B bi-subunit; coupling the artificial gene with the gene sequence of the cholera toxin B subunit to form a fusion gene;and expressing the fusion gene by using an escherichia coli prokaryotic expression system, and performing protein purification to obtain the helicobacter pylori multiepitope vaccine. The helicobacterpylori multiepitope vaccine can induce an organism to generate T cell immune response and high-titred specific antibody humoral immune response aiming at the urease A and B bi-subunit, and can be used for preventing and treating helicobacter pylori infection related diseases.

The invention discloses a Bacillus amyloliquefaciens WH3 strain, and a preparation method and application thereof. The preparation method comprises the following steps: 1, separation and identification of bacteria: separating bacteria resistant to rape sclerotinia rot from rape seedlings, and carrying out 16SrDNA and morphological identification to determine that the WH3 strain obtained by separation is Bacillus amyloliquefaciens; 2, separation, purification and identification of an antifungal active substance Safenour: fermenting the WH3 strain, extracting the antifungal active substance, separating and purifying through a sephadex column, and carrying out MALDI-TOF (matrix-assisted laser desorption/ionization-time of flight ) mass spectrometry on the active antifungal substance to inferthat the substance is a ring type polypeptide; and 3, application of the strain in the preparation of vaccines and immunological adjuvants. After the Safenour used as the adjuvant is mixed with a protein antigen and mice are respectively immunized through oral administration and injection of the mixture, effective body fluid and cellular immune response can be activated, and a high-titer specificantibody can be detected in the blood serum. The Safenour has low production cost and high stability, does not need to be emulsified when mixed with the antigen, and has a better immunoenhancement effect in comparison with Freund adjuvants and cholera toxin B subunits.

The invention discloses a multimeric protein having the effect of brain targeting, and a preparation method and usage thereof. The method comprises the steps: firstly, expressing cholera toxin B subunit and a fusion protein EGFP-CTA2-TAT of three proteins: an enhanced green fluorescent protein, a cholera toxin A subunit and cell-penetrating peptide in escherichia coli by incompatible double plasmid systems to obtain CTB gene by PCR amplification; cloning the gene segment into a carrier pET-28a to obtain a recombinant plasmid pET-28a-CTB; using wild type CTA2, EGFP and TAT amino acid sequences as templates, inserting 3 enzyme cutting sites and linkers between EGFP and CTA2, and optimizing to obtain codons suitable for expression in escherichia coli; cloning the gene segment into a carrier PET-22b (+) to obtain recombinant plasmid PET-22b-EGFP-CTA2-TAT; using different resistances of PET-28a-CTB and PET-22b-EGFP-CTA2-TAT, and co-transforming the different resistances of the PET-28a-CTB and the PET-22b-EGFP-CTA2-TAT into escherichia coli BL21, to obtain engineering bacteria after screening under double resistance selection pressure of penbritin and kanamycin. CTB5/EGFP-CTA2-TAT chimeric proteins can be obtained after inducible expression of the engineering bacteria by IPTG. The invention further discloses the preparation method and usage of the protein.

The invention discloses a meningococcus capsular polysaccharide polyvalent multivalent conjugate vaccine, a preparation method and application thereof. The polyvalent multivalent conjugate vaccine comprises at least two conjugates of group A, group C, group Y or Group W135 meningococcus capsular polysaccharide and cholera toxin B-subunit. The invention improves the method for extracting the meningococcus capsular polysaccharide, and improves the yield and purity of the capsular polysaccharide by adopting a flocculation extraction method under an electric field.

The invention provides a helicobacter pylori epitope vaccine. The active constituent of the helicobacter pylori epitope vaccine is a polypeptide and mainly consists of a cholera toxin B subunit and a B cell antigen epitope from a urease A subunit. A preparation method of the helicobacter pylori epitope vaccine comprises the following steps of: synthesizing the nucleotide sequence of the B cell antigen epitope from the urease A subunit by a PCR (polymerase chain reaction) technology, coupling the nucleotide sequence with the gene sequence of the cholera toxin B subunit to form into a fusion gene, and expressing the fusion gene in the escherichia coli by an expression vector to obtain the fusion protein of the epitope vaccine by means of protein purification. The helicobacter pylori epitope vaccine can be used for inducing the human body to generate an epitope specific antibody which is higher in titer to the urease. In the biologic medical field, the epitope vaccine can be used for preventing and curing the relevant diseases caused by the helicobacter pylori infection, thereby being great in economic benefit and social benefit.

The invention relates to preparation and application of a mycoplasma hyopneumoniae multi-epitope mucosal vaccine. A mycoplasma hyopneumoniae membrane protein, an adhesive protein P97, a lipoprotein P65, a specific membrane protein P46, a B cell epitope, a Th epitope, a CTL epitope and a cholera toxin subunit B are taken as a vaccine frame structure, a pRSETA carrier is cloned in through flexible linker connection, then Escherichia coli is transformed, and fermentation, purification and preparation technologies are carried out, so that the mycoplasma hyopneumoniae multi-epitope mucosal vaccine with ideal immunogenicity is obtained. A self-made mucosal adjuvant is used in a preparation process, so that production and using processes of the vaccine are simpler and more convenient. Animal experiments show that the mycoplasma hyopneumoniae multi-epitope mucosal vaccine not only has good safety but also can stimulate effective mucosal immunity, humoral immunity and cellular immune reactions.

The invention discloses a vaccine for blocking transmission of echinococcosis pathogeny echinococcus granulosus from a source. The vaccine provided by the invention is a CTB-EgM123 vaccine or a complete set of vaccine consisting of the CTB-EgM123 vaccine and a subunit vaccine of an EgM123 protein recombinant mycobacterium smegmatis vaccine. An active ingredient of the CTB-EgM123 vaccine is a fusion protein formed by fusion of a cholera toxin B subunit and an echinococcus granulosus EgM123 protein; the active ingredient of the EgM123 protein recombinant mycobacterium smegmatis vaccine can express the recombinant mycobacterium smegmatis of the echinococcus granulosus EgM123 protein. The vaccine provided by the invention can stimulate the immune response in a host, improves the immune level of the host, enhances the intestinal mucosal immune response, and plays an important protective role in resisting infection of echinococcosis.

The invention discloses transformant. The transformant comprises a recipient bacterium and a recombinant vector transferred to the recipient bacterium, wherein the recipient bacterium is lactococcus lactis NZ9800; the original vector of the recombinant vector is pNZ8112; and the exogenous target gene of the recombinant vector comprises a porcine somatostatin gene, a cholera toxin B subunit gene and a cell wall anchor stator M6 gene. The transformant can enter the porcine intestinal tract by oral administration; compared with injection immunization, the oral immunization mode reduces the toxicity caused by direct contact with a circulatory system and increases the safety of vaccines; and the immune response of organisms is enhanced and the immune effect is improved by combined immunization of the transformant, an antigen gene and an immune enhancing factor.

A process for preparing cholera toxin B subunit features that by the expression system of Bombyx mori nuclear polyhedrosis virus, the virus with CTB gene is recombined to obtain the recombinant virus (CGMCC No.0594) which has been preserved by Ordinory Microbe Preservation Center of China Microbial Strain Preservation and Management Committee. An oral medicine for treating cholera is prepared through inoculating the virus to the larva or pupa of Bombyx mori, expressing, separating, purifying, and freeze drying. Its advantages are high curative effect, and low cost.

The invention discloses a fusion protein vaccine capable of inhibiting Streptococcus and/or preventing Streptococcus infection. With fusion protein obtained through linkage of sortase A and a cholera toxin B subunit through connecting peptide, the Th17 cell activation level is remarkably increased, the effects of preventing pathogenic bacteria from settling and quickly removing the pathogenic bacteria can be realized, and the fusion protein has protecting effects on Group A Streptococcus of different serotypes and has the superiority of high efficiency, broad spectrum and low cost. With the fusion protein, the use amount of immunogen is reduced, the production technology is simplified, and the production cost is reduced. Meanwhile, the vaccine adopts the way of mucosal immunity, has the characteristics of being free of tissue damage, free of local side effects and convenient to use, and is easy to popularize and use.

The invention provides an Asia 1-type FMDV compound multiepitope which adopts an Asia 1-type FMDV capsid protein VP1 antigen activity area as the basic skeleton, and comprises the epitope of Asia 1/JL/05 plant VP1, the epitope of the IND 63/72 plant VP1, the epitope of YNBS/58 plant VP1, cholera toxin B subunit and helper T lymphocyte epitope. The compound multiepitope can be used for preparing Asia 1-type FMDV subunit vaccine, nucleic acid vaccine, synthetic peptide vaccine, live vector vaccine and a plurality of genetic engineering vaccines.

The invention relates to an immune modulator for treating arteriosclerosis. Aiming at antigen peptides's weak immunogenicity, a series of epitopes peptides of antigens (SEQ ID NO.1) originating from heat shock protein (HSP60) are inserted into the downstream of Cholera toxin B subunit(CTB), and fusion expression of antigens epitopes peptides and CTB is realized. Produced fusion protein (CTB-p277) and Heat shock protein 65 (HSP65) are coupled chemically, without adjuvant, and directly used immunization. The coupled protein can stimulate body to produce high titer antibody aiming at P277 through mucosal immunity pathway, obviously inhibit occurrence of New Zealand white rabbits's arteriosclerosis, and can have a function for treating arteriosclerosis.

The present invention discloses a preparation method of cholera toxin B subunit protein having biological activity. According to the present invention, a lot of cholera toxin subunits are expressed in Escherichia coli by using a prokaryotic expression vector, and then in vitro efficient renaturation is performed to obtain a large number of the cholera toxin B subunit having biological activity; the operations can be performed in the general laboratory, and no pollution is generated; and the most important advantage is that the labeling with various fluorescence, isotopes and tracers can be performed according to the experimental needs.

The invention provides a novel fasciola hepatica multi-epitope vaccine. An active ingredient of the novel fasciola hepatica multi-epitope vaccine is a polypeptide. The polypeptide is mainly formed by fasciola hepatica sphingolipid activated mucin-like protein-2 Th, a multicopy body of B-cell epitope and a mucosal immunologic adjuvant cholera toxin B subunit. An artificial gene is mainly synthesized through the gene synthesis technology and comprises Th of sphingolipid activated mucin-like protein-2 and a gene sequence of the multicopy body of the B-cell epitope, then the artificial gene is coupled with the gene sequence of the cholera toxin B subunit, and a fusion gene is formed. An escherichia coli prokaryotic expression system is utilized for expressing the fusion gene, and the fasciola hepatica multi-epitope vaccine is obtained after protein purification. The fasciola hepatica multi-epitope vaccine can induce an organism to generate sphingolipid activated protein T cell immune responses and high-titer specific antibody humoral immune responses, and can be used for preventing and treating fasciola hepatica infection related diseases.

The invention discloses a vibrio cholerae O139 capsular polysaccharide conjugate vaccine which comprises a conjugate of vibrio cholerae O139 capsular polysaccharide and a protein carrier, wherein the protein carrier is preferential to be a cholera toxin B-subunit. The invention also provides a preparation method of the conjugate vaccine, which comprises the step of coupling the vibrio cholerae O139 capsular polysaccharide and the carrier protein with a mass ratio of (0.2-3):1. After the vibrio cholerae O139 capsular polysaccharide conjugate vaccine is subcutaneously injected to an immune mouse, the conjugate vaccine can excite the mouse to generate antibodies for resisting vibrio cholerae O139 and antibodies for resisting cholera toxin, thus the titer of vibriocidal antibodies can be improved by more than 2 times, and the titer of the antibodies for resisting the cholera toxin (IgG and IgA) can be improved by more than 4 times.

The invention discloses a polysaccharide and protein conjugate. The polysaccharide and protein conjugate is formed by coupling vibrio cholerae lipopolysaccharide and cholera toxin non-toxic subunit protein, wherein the vibrio cholerae lipopolysaccharide is from vibrio cholerae 0139, the protein content in the vibrio cholerae lipopolysaccharide is not more than 2 percent, the nucleic acid content is not more than 5 percent, and the molecular weight of the vibrio cholerae lipopolysaccharide is 8Kd. The immune effect of the polysaccharide and protein conjugate is higher than that of common rBS-WC (recombinant B subunit-killed whole-cell cholera) vaccine, single LPS (lipopolysaccharide) vaccine or single CTB (cholera toxin B-subunit) vaccine, the conjugate can enhance the immunogenicity of LPS to make an induced immunity reaction have immunologic memory and potentiation effect to generate massive bactericidin, is suitable for various kinds of people, and can initiate mucosal immunity of a body to realize IgA (immunoglobulin A) antibodies massively exist in intestinal secretion fluid and bile.

The invention provides a novel alveolar echinococcosis subunit vaccine. A polypeptide serves as an active ingredient of the alveolar echinococcosis subunit vaccine, which is mainly composed of a surface antigen Emy162 of alveolar echinococcosis and a mucosal immune adjuvant, namely cholera toxin B subunit (CTB). According to the invention, the gene sequence of the alveolar echinococcosis surface antigen Emy162 is synthesized by virtue of a gene synthesis technology, and then the gene is coupled with the gene sequence of the cholera toxin B subunit, so that a fusion gene is formed. The fusion gene is expressed by virtue of an escherichia coli prokaryotic expression system, and protein is purified, so that the alveolar echinococcosis subunit vaccine is obtained. The alveolar echinococcosis subunit vaccine is capable of inducing a body to generate alveolar echinococcosis targeted T cell and B cell immune response and high-titer specific antibody humoral immune response, and the subunit vaccine can be used for preventing and treating alveolar echinococcosis infection related diseases.

The invention discloses an expression vector PVX-6His-CTBt-Bt for producing a multi-epitope vaccine of hepatitis c virus. The multi-epitope vaccine 6His-CTBt-Bt of hepatitis c virus expressed by the vector comprises a histidine tag, an orally-taken immunologic adjuvant cholera toxin B subunit (CTB) and six B cellular antigen epitopes Bp of refractory hepatitis c virus genetype 1. To facilitate high-efficiency expression of the vaccine in tobacco, the 6His-CTBt-Bt gene optimized by an artificially synthesized codon is cloned to a high-efficiency plant virus expression vector potato X virus vector PVX201 to obtain the recombinant expression vector PVX-6His-CTBt-Bt of the multi-epitope vaccine 6His-CTBt-Bt of the hepatitis c virus; the expression vector is inoculated to the tobacco; the tobacco is regarded as a reactor to produce the anti-hepatitis c virus vaccine which aims to the refractory hepatitis c virus, is easy to purify, and can achieve a good orally-taken immune effect and resist against virus infection in vitro.