A synthetic gonadotropin-releasing hormone (GnRH) vaccine for control of fertility and hormone dependent diseases without any adjuvant.

Active immunization against self-peptides have gained widespread acceptance inspite of their low immunogenicity. Recent applications involving multiple copies of self-peptides in linear alignment and conjugation with carrier proteins appear to increase the immune response against self-peptides. As with most vaccines, however, immunogens require supplementation with adjuvants to elicit an optimum immune response. In the present study, we prepared a double-chain mini-protein with each chain containing three linear repeats of the self-peptide gonadotropin-releasing hormone (GnRH3), the hinge region of human IgG1 (hinge), and a T-helper epitope from the measles virus protein (MVP). The GnRH3-hinge-MVP mini-protein was conjugated to purified recombinant heat shock protein 65 (Hsp 65) of Mycobacterium bovis and used to immunize rats primed with subcutaneous injections of Bacillus Calmette-Guerin (BCG) in the absence of adjuvants. The GnRH3-hinge-MVP-Hsp 65 stimulated the production of specific anti-GnRH antibodies in the absence of adjuvants and the antibody titer was comparable to that produced in rats immunized with the dimeric mini-protein in the presence of Freund's adjuvant. Moreover, immunization with the adjuvant-free GnRH3-hinge-MVP-Hsp 65 induced degeneration of the reproductive organs in both male and female rats unlike those immunized in the absence of Hsp 65 or in control animals inoculated with the vehicle only. Histological examination of the affected organs showed atrophy of the seminiferous tubules with diminished spermatogenesis in the testes of male rats. In female rats, the uteri were much smaller in size and the ovaries exhibited reduced follicular development. These findings demonstrated that GnRH3-hinge-MVP-Hsp 65 mounted a strong immune response in the absence of conventional adjuvants, and could prove useful in control of fertility and the treatment of conditions/diseases where GnRH ablation is required.

The immunogenicity of recombinant and dimeric gonadotrophin-releasing hormone vaccines incorporating a T-helper epitope and GnRH or repeated GnRH units.

In this study, we designed two linear peptides, GnRH-hinge-MVP, which consists of human gonadotrophin-releasing hormone (GnRH), hinge fragment 225-232/225'-232' of human IgG1 and a T helper peptide from measles virus protein (MVP), and GnRH3-hinge-MVP, which contains three copies of GnRH (so termed GnRH3). The DNA constructs encoding for the two peptides were fused to the C-terminal encoding sequence of asparaginase, encompassing residues 199-326, through an acid-labile aspartyl-prolyl linker. The chimeric genes were expressed at high levels in Escherichia coli. The fusion proteins were purified to approximate homogeneity by means of washing the inclusion bodies and by ethanol precipitation. The GnRH-hinge-MVP or the GnRH3-hinge-MVP was released from the fusion proteins by cleavage with hydrochloric acid and further oxidized into double-chain miniproteins after purification. Both dimeric constructs proved to be efficient immunogens. It was shown that rats immunized with the immunogens generated antibodies specific for GnRH. The dimeric GnRH3-hinge-MVP containing three copies of GnRH in each chain induced a higher titre of anti-GnRH antibodies than the GnRH-hinge-MVP, containing a single copy of GnRH in each chain. These results demonstrate that combining multicopies or single copies of peptide with hinge fragment of human IgG and T helper peptide from measles virus protein can induce anti-peptide immune responses. Our data also suggest that these methods of preparation and dimerization of the recombinant polypeptides may provide a useful strategy for other polypeptide vaccine developments.

Study on Preparation and Identification of Immobilized Metal Ion Affinity Adsorption Gel / 生物化学与生物物理进展

Sepharose 6B was activated by epichlorohydrin in the strong base condition, and then reacted with solution of iminodiacetic sodium. The arms of IDA were conjuncted to the activated Sepharose 6B. Then the products were reacted with the solution of NiSO4. The arms of IDA were chelated with Ni2+,and the chelating resin―Ni2+-IDA could be prepared. The physicochemical indexes and performance in purifying protein of the expressing product were assayed with atomic absorption method and purifying aimed protein-human B lymphocyte stimulator(hBLyS) from the expressing products in E.coli. The results indicated that the performance of made gel is very good, and its price is less than 1/10 of that of commodity gel.

THE ANTIBACTERIAL PEPTIDE CM4 INHIBITS REGENERATION OF SACCHAROMYCES CEREVISIAE PROTOPLAST BY DAMAGING ON THE PLASMA MEMBRANES / 微生物学通报

The antibacterial peptide CM4 having potent antifungal activity on inhibitiong the cell wall regeneration of Saccharomyces cerevisiae protoplasts.When the peptide increased,the ratio of the regenerated colonies drop obviously.To study the antifungal mechanism of the antibacterial peptide,fluorescence\|labeled peptide mixted with the protoplast of yeast,then confocal laser scanning microscopy were performed.The results indicated that the peptides interactted with the protoplast membrane and damaged the structure of the membrane,then the permeation of protoplast changed.Finally the protoplasts with the peptide failed to regenerate the cell walls leading to killing the cell.