MXPA00001706A - Fusion proteins comprising carriers that can induce a dual immune response - Google Patents

Abstract

The subject invention provides a fusion protein for producing a dual immune response in a vertebrate, which fusion protein comprises:(a) a first proteinaceous portion analogous to all or part of a peptide endogenously synthesized within the vertebrate, the activity of which peptide is to be inhibited within the vertebrate, and which proteinaceous portion by itself is incapable of eliciting an effective immunoinhibitory response in said vertebrate;connected to (b) a second proteinaceous portion analogous to all or part of an immunogen from a pathogen, which pathogen is capable of pathogenically infecting the vertebrate;the portion (b) causing the vertebrate's immune system to recognize the portion (a) and produce a response that:(i) inhibits the activity of the peptide endogenously synthesized within the vertebrate;and (ii) protects the vertebrate from infectionby the pathogen, when the vertebrate is vaccinated with an effective amount of the fusion protein. The subject invention also provides fusion proteins which comprise a proteinaceous portion (b) that is a carrier that is analogous to all or part of a BHV-1 antigen, which fusion proteins induce in a vertebrate vaccinated with an effective amount of such fusion protein an immune response that inhibits the activity of a peptide as recited in (a), above.

Description

FUSION PROTEINS THAT COMPRISE SUPPORTS THAT CAN INDUCE A DOUBLE IMMUNE RESPONSE FIELD OF THE INVENTION The present invention is in the field of animal and human health, and is directed to fusion proteins that are useful in vaccine compositions.

BACKGROUND OF THE INVENTION The vertebrate immune system comprises an intricate system of cells, secreted factors and responses to protect an organism with respect to a pathogenic infection by microbes, viruses, toxins, and other pathogens and irritants. Certain molecules, however, comprise epitopes that do not induce an effective immune response in a vertebrate because of their small size and / or since they are synthesized endogenously within this vertebrate and therefore are not perceived as "foreign" by the system. immune of the vertebrate. Methods for producing antibodies against certain peptides that are normally non-immunogenic, such as hormones, are desirable since it can thereby achieve immune regulation of the activity of said peptides within the organism. Various peptide hormones have been combined with various support peptides in fusion proteins to produce an effective immune response against the hormone when an organism is vaccinated with the fusion protein. The support portion causes the body's immune system to recognize and generate antibodies against the hormonal peptide that it would otherwise not generate. U.S. Pat. 5,403,586 issued to Russell-Jones et al., For example, refers to fusion proteins comprising a protein similar to gonadotropin-releasing hormone (GnRH), also known as luteinizing hormone-releasing hormone (LHRH), and an analogous protein to TraTp, in which the presence of the analogue to TraTp in the fusion protein helps to trigger the production of anti-GnRH antibodies. TraTp is an outer membrane lipoprotein produced by certain strains of E. coli, such as described in US Pat. 5,403,586, noted above. U.S. Pat. 5,422,110 issued to Potter et al., Refers to support systems that include chimeric proteins comprising a leukotoxin polypeptide fused to a selected antigen. Leukotoxin works to increase the immunogenicity of the antigen. Selected antigens described in the present context include GnRH, somatostatin (SRIF), and rotavirus viral protein 4 (VP4). PCT Patent Document WO 90/02187 relates to fusion proteins comprising an antigenic hydrophilic portion, such as hepatitis B surface antigen (HBsAg, Hepatitis B surface antigen), and a peptide, such as GnRH , which by itself is not substantially antigenic. GnRH is a decapeptide produced endogenously, mainly in the hypothalamus. It is preserved to a high degree among vertebrate species. In mammals, the GnRH gene encodes the decapeptide glu-his-trp-ser-tyr-gly-leu-arg-pro-gly with subsequent post-translational modification of the terminal ends of N and C with pyroglutamic acid and glycinamide , respectively, producing (pyro) -glu-his-trp-ser-tyr-gly-leu-arg-pro-gly-NH2- It has been shown that GnRH plays a critical role in the regulation of reproductive functions in all vertebrates elderly, regulating the production and release of the follicle-stimulating hormone (follicle-stimulating hormone FSH) and luteinizing hormone (luteinizing hormone-LH) from the pituitary gland. Since FSH and LH play a certain role in spermatogenesis and ovulation, as well as in steroidogenesis, vaccines that result in the production of antibodies against GnRH lead to the suppression of reproductive function (fertility) in both males as in females, and should also control secondary sexual characteristics such as behavior related to sex. In males, LH regulates steroidogenesis in Leydig cells. Therefore, active immunization of males against GnRH leads to testicular atrophy and a decrease in testosterone production and testicular function (Ladd, A. et al., 1994, Biol. Reprod. 51: 1.076-1.083 Ladd A., 1993, Am. J. Reprod. Immunol. 29: 189-194).

A vaccine against GnRH has been approved by the United States Food and Drug Administration as a new drug under investigation for the treatment of prostate cancer (Ladd A., 1993 , previous appointment). The development of an immune contraceptive with GnRH is a useful alternative to surgical sterilization in animals, and has the added advantage of being reversible, since spermatogenesis and fertility can return to normal simply by reducing the anti-inflammatory titers. GnRH (Ladd A. et al., 1989, J. Reprod. Immunol. 15: 85-101). However, since GnRH is a small self-peptide and has a short half-life (WO 90/02187, March 8, 1990) it is only weakly immunogenic, even though it is injected with a potent adjuvant. For example, a significant proportion of animals are not able to mount an effective antibody response against GnRH when it is administered in Freund's complete adjuvant. In order to generate an important antibody response, GnRH must therefore be conjugated, either chemically or recombinantly, with a supporting protein. None of the aforementioned references, however, teach or suggest using a support that triggers an immune suppressive response against itself.

NOVELTY OF THE INVENTION The present invention provides a fusion protein for producing a double immune response in a vertebrate, which fusion protein comprises: (a) a first protein portion analogous to all or a portion of a peptide endogenously synthesized within the vertebrate, the activity whose peptide must be inhibited within the vertebrate, and whose protein portion is itself incapable of producing an effective immune inhibitory response in said vertebrate; connected with (b) a second protein portion analogous to all or a part of an immunogen from a pathogen, the pathogen of which is capable of infecting the vertebrate pathogenically; giving rise to portion (b) that the vertebrate immune system recognizes the portion (a) and produces a response that: (i) inhibits the activity of the peptide synthesized endogenously within the vertebrate; and (ii) protects the vertebrate with respect to an infection by the pathogen, when the vertebrate is vaccinated with an effective amount of the fusion protein. The present invention further provides, in a second aspect, a fusion protein for producing an immune response in a vertebrate, which fusion protein comprises: (a) a first protein portion analogous to all or a portion of a peptide, the activity whose peptide must be inhibited within the vertebrate, and whose protein portion is itself incapable of producing an effective immune inhibitory response in said vertebrate; connected with (b) a second protein portion analogous to all or a part of a bovine herpesvirus type-1 antigen (Bovine Herpes Virus Type-1 BHV-1); the second protein portion (b) giving rise to the vertebrate's immune system recognizing the first protein portion (a) and producing an immune response capable of inhibiting the activity of the peptide within the vertebrate when the vertebrate is vaccinated with an effective amount of the fusion protein. The present invention further provides fusion proteins, as cited in the preceding two paragraphs, which are recombinant fusion proteins. The present invention further provides a polynucleotide molecule comprising a nucleotide sequence encoding a fusion protein of the present invention. The present invention further provides a vector comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding a fusion protein of the present invention. The present invention further provides a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding a fusion protein of the present invention. The present invention further provides a double-function vaccine comprising a certain amount of a fusion protein, as set forth above, comprising: (a) a first protein portion analogous to all or a portion of a peptide endogenously synthesized within a vertebrate, the activity of which peptide is to be inhibited within the vertebrate, and whose protein portion is itself incapable of producing an effective immune inhibitory response in said vertebrate, connected with (b) a second protein portion analogous to all or a part of an immunogen from a pathogen, whose pathogen is capable of infecting the vertebrate pathogenically; wherein portion (b) is capable of causing the vertebrate immune system to recognize portion (a) and produce a response that: (i) inhibits the activity of the peptide synthesized endogenously within the vertebrate; and (ii) protects the vertebrate with respect to an infection by the pathogen, said fusion protein being present in the double-function vaccine in an amount effective to inhibit the activity of the peptide from which the portion (a) is derived and to protect the vertebrate with respect to an infection by the pathogen from which the portion (b) is derived, said double-acting vaccine also comprising a support (vehicle) acceptable for pharmaceutical or veterinary use. The present invention further provides a method for inhibiting the activity of a peptide synthesized endogenously in a vertebrate and for protecting the vertebrate with respect to a pathogenic infection, which method comprises immunizing the vertebrate with a vaccine as cited in the preceding paragraph in a effective amount to inhibit the activity of the peptide and to protect it against an infection by the pathogen. The present invention further provides a vaccine for inhibiting the activity of a peptide in a vertebrate, comprising a fusion protein as set forth above, comprising: (a) a first protein portion analogous to all or a portion of a peptide, the activity of which peptide is to be inhibited within the vertebrate, and whose protein portion is itself incapable of producing an effective immune inhibitory response in said vertebrate; connected with (b) a second protein portion analogous to all or a part of a BHV-1 antigen; the second protein portion (b) being capable of causing the vertebrate immune system to recognize the first protein portion (a) and produce a response that inhibits the activity of the peptide within the vertebrate, the fusion protein being present in the vaccine in an amount effective to inhibit the activity of the peptide in the vertebrate, and the vaccine further comprising a carrier (vehicle) acceptable for pharmaceutical or veterinary use. The present invention further provides a method for inhibiting the activity of a peptide in a vertebrate, comprising immunizing the vertebrate with a vaccine as cited in the preceding paragraph in an amount effective to inhibit the peptide. The present invention further provides a method for producing polyclonal antibodies directed against a peptide that is endogenously synthesized in a vertebrate, comprising vaccinating said vertebrate with an antibody-inducing amount of a fusion protein of the present invention, or of a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, which fusion protein comprises a portion (a) analogous to all or a portion of a peptide synthesized endogenously within the vertebrate; obtain a serum containing polyclonal antibodies from the vaccinated vertebrate; and isolating polyclonal antibodies from the serum that bind to the endogenously synthesized peptide; thereby producing polyclonal antibodies directed against the peptide. The present invention further provides polyclonal antibodies directed against a peptide synthesized endogenously produced according to the method recited in the preceding paragraph. The present invention further provides a method for producing a monoclonal antibody directed against a peptide that is endogenously synthesized in a vertebrate, comprising vaccinating said vertebrate with an antibody-inducing amount of a fusion protein of the present invention, or of a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, which fusion protein comprises a portion (a) analogous to all or a portion of a peptide synthesized endogenously within the vertebrate; and isolating a spleen cell from the vaccinated vertebrate, whose spleen cell excretes a monoclonal antibody that binds specifically to the endogenously synthesized peptide; thus producing a monoclonal antibody directed against the peptide. The present invention further provides monoclonal antibodies directed against an endogenously synthesized peptide, produced according to the method cited in the preceding paragraph.

BRIEF DESCRIPTION OF THE FIGURES Figure 1: Artificial structures (constructions) of gD and GnRH fusions: fusion proteins constructed in accordance with the present invention are described. The gD in these artificial structures is mature (the signal sequence has been removed) and truncated (the transmembrane domain and the remaining 3 'sequence have been removed). GnRH is in the tetramer form. Figure 2: A clone of the GnRH tetramer, constructed by fusing the C terminal ends of the reshaped GnRH oligonucleotides (set forth in SEQ ID NOS: 7 and 8) with the GnRH sequence (annealed oligonucleotides set forth in SEQ ID NOS: 9 and 10) in the dimer clone of 98BS and GnRH. Flanking sequences from plasmid pBS KS + (Stratagene) and from cloning sites existing therein are also described. This nucleotide sequence is set forth in SEQ ID NO: 14. The encoded amino acid sequence is also shown (SEQ ID NO: 15). Figure 3 (Fig. 3A-3C): The nucleotide sequence (SEQ ID NO: 16) encoding the gD of BHV-1 within the clone FlgD / Pots207nco (# 79) as well as the coding sequence of the poly (amino acid) gD (SEQ ID NO: 17). Nucleotides 3 56 encode the signal sequence; nucleotides 1,092-1,169 encode the transmembrane domain (tmgD). Nucleotides 57-1,259 encode mature gD, and nucleotides 57-1,076 encode truncated mature gD. "Gly" represents regions of glycosylation. Shown are vector sequences flanking the sequence encoding gD. Figure 4 (Fig. 4A-4C): Alignment report (DNA alignment) comparing gD of BHV-1 from clone FlgD / Pots207nco (# 79) (gD / Pots, top sequence) and gD of BHV- 1 having accession number M59846 to the Gene Bank (GenBank) (bottom sequence) (Tikoo et al., 1990, cited above) (GenBank DNA sequence database of the US National Center for Information Biotechnology (US National Center for Biotechnology Information, (NCBI, Bethesda, Maryland)) The Clustal method with weighted residual weights table was used for this report. "TM" represents a transmembrane domain, the residues pigeonholed in the FlgD clone. / Pots207nco (# 79) are the ones that differ from the residues in M59846. The DNA of M59846 is SEQ ID NO: 18. Figure 5: Alignment of amino acids between gD / Pots (lower sequence) and M59846 (sequence superior) The Clustal method was used with a PAM250 residue weight table. The residues in the gD / Pots that differ from the residues in the M59846 are pigeonholed. M59846 is SEQ ID NO: 19. Figure 6 (Fig. 6A-6C): pQE-tmgD. The sequence encoding nucleotides for tmgD, flanked by sequences of plasmid pQE-31, including a sequence encoding a 6XHIS tag, which is expressed in the state connected to the tmgD (SEQ ID NO: 20). The amino acid sequence of the tmgD is also shown with the 6XHIS tag connected (SEQ ID NO: 21). Figure 7 (Fig. 7A-7C): Sequence encoding nucleotides and flanking sequences for the plasmid pQE-GnRH: gD (SEQ ID NO: 22). Also shown is the amino acid sequence of the fusion protein 4GnRH-tmgD, which includes a 6XHIS tag (SEQ ID NO: 23). Figure 8 (Fig. 8A-8C): pQE-gD: GnRH. The sequence encoding nucleotides and flanking sequences of the plasmid are shown (SEQ ID NO: 24). The amino acid sequence of tmgD-4GnRH is also shown, with a 6XHIS tag (SEQ ID NO: 25). Figure 9 (Fig. 9A-9C): pQE-GnRH: gD: GnRH. The sequence encoding nucleotides and plasmid flanking sequences are shown (SEQ ID NO: 26). Also shown is the amino acid sequence of 4GnRH-tmgD-4GnRH with a 6XHIS tag (SEQ ID NO: 27). Figure 10: Comparison of expression products from artificial structures of the bacterial vector pQE. "A" is pQE-tmgD, "B" is pQE-gD: GnRH, "C" is pQE-GnRH: gD, and "D" is pQE-GnRH: gD: GnRH. The amino acids that bind to the gD portions, the GnRH tetramers, and the 6H XIS labels are reproduced in this figure. Figure 1 1 (Fig. 1 1 A-1 1 B): Nucleotide sequence (SEQ ID NO: 28) from the pCMV-tgD plasmid encoding a truncated gD, and deduced amino acid sequence (SEQ ID NO: 29) of the truncated gD expression product including the signal sequence.

Figure 12 (Fig. 12A-12B): Nucleotide sequence (SEQ ID NO: 30) from the plasmid pCMV-gD: GnRH (Accession No. 203370 ATCC) encoding a tgD-4GnRH fusion protein, with the sequence deduced from amino acids (SEQ ID NO: 31) of the fusion protein product including the signal sequence.

DETAILED DESCRIPTION OF THE INVENTION Fusion Proteins In a first aspect, the present invention provides fusion proteins that induce in a vertebrate a double immune response that inhibits the activity of a peptide endogenously synthesized by the vertebrate and also inhibits a pathogenic infection in the vertebrate. The inhibition of the endogenously synthesized peptide is obtained by connecting a first protein portion, which is analogous to all or a portion of the endogenously synthesized peptide, to a support, the support being a protein portion analogous to all or a portion of an immunogen from a pathogen capable of infecting the vertebrate pathogenically. In addition to functioning as a support (i.e., enhancing an immune response against the protein analogous to the endogenously synthesized peptide or a portion thereof), the analogous portion to the immunogen or to a part of the immunogen from the pathogen also induces a response against itself in the vertebrate and therefore protects the vertebrate with respect to infection by the pathogen.

Since two major causes of economic loss in large-scale feedlot cattle (feedlot) in the United States of America and in range-fed cattle globally are bovine respiratory disease (BRD from Bovine Respiratory Disease) caused by an infection by BHV-1 and a sexual and aggressive behavior, a product that simultaneously treated the BRD and also inhibits the secondary sexual characteristics, for example, of aggression, would improve the quality and the productivity of the skeletons eliminating or reducing these infectious and endocrine causes of production losses in cattle. Therefore, as an embodiment of the present invention, the glycoprotein D (gD), which is an immunogenic agent from BHV-1, was selected as a support to be combined with a GnRH peptide in a fusion protein with the to regulate GnRH activity in cattle while simultaneously providing protection against the BRD. Certain protein portions that are analogous to a immunogen from a pathogen or to a part of an immunogen from a pathogen, such as those analogous to the BHV-1 glycoprotein described herein, have not been previously described or suggested as carriers. . Therefore, a second aspect of the present invention provides a fusion protein for producing an immune response in a vertebrate, whose fusion protein comprises as a support a protein portion analogous to all or a portion of an antigen of BHV 1. In this second aspect, the vertebrate does not need to be a vertebrate that is capable of being infected pathogenically by BHV-1; the protein analogous to BHV-1 antigen acts simply as a support that induces an immune response that inhibits the activity of the protein portion (a). Therefore, if a fusion protein of this invention comprises, for example, a portion (a) analogous to all or a part of a GnRH peptide, and a portion (b) analogous to all or a portion of a BHV 1 antigen, said fusion protein will produce a double immune response in cattle, but it will also be useful in other vertebrate species to inhibit the activity of GnRH without protecting against an infection with BHV-1, since these other species are not pathogenically infected by BHV-1. For the purposes of this invention"fusion protein" means a molecule comprising a plurality of protein portions connected together. Thus, the fusion proteins of this invention include chemical conjugates (portion (a) and portion (b) chemically connected) and recombinant fusion proteins. A fusion protein according to this invention comprises a protein portion (a) and a protein portion (b), whereby it is understood that the molecule can comprise at least a portion (a) and at least a portion (b) ), but may comprise more than one portion (a) and / or more than one portion (b). Portions (a) and (b) can be connected linearly. If multiple portions of (a) and / or (b) are present, then the portions may be connected linearly or may be connected in a branched manner, for example with one of the portions (a) or (b) centrally placed in the molecule and with other portions (a) or (b) connected multiplely to the central portion. Other connection patterns, which can be determined by those with ordinary experience in the technology sector, are included within the present invention, provided that at least a portion (a) and at least a portion (b) ) are present in the fusion proteins. Portions (a) and (b) may be positioned with respect to one another such that an effective immune response against portion (a) as well as against portion (b) is optimized if desired. Such placement can be determined by methods known to those with ordinary experience in the technology sector. For example, a fusion protein as described herein can be tested by Western blotting with antibodies against (a) and / or (b) to determine whether portions (a) and / or (b) are positioned in a manner such that the fixation of specific antibodies for them is optimized. The portions of the fusion proteins in question can be connected by means that include chemical connections and recombinant connections. A "chemical connection" involves creating a chemical intermediate compound from a protein portion, and reacting this intermediate compound with another protein portion. For example, a "chemical connection" can involve the formation of a direct covalent bond between an organic group of a protein portion, such as portion (a), and an organic group of the other protein portion, e.g. portion (b), with the proviso that the portions must have organic groups that are capable of reacting under appropriate reaction conditions to form said covalent bond. As another example, one of the protein portions, such as portion (a), can be derivatized to form an intermediate compound containing substituents that react with portions (b). A "recombinant connection" involves ligating a nucleic acid encoding a protein portion with a nucleic acid encoding another protein portion, and expressing a protein therefrom in an appropriate expression system. Chemical connections and a recombinant connection are known in the art and are described in greater detail in the present text. The term "support" as used in the present text (except when it appears in the phrases "pharmaceutically acceptable carrier (vehicle)", "support (vehicle) acceptable for pharmaceutical or veterinary use" and similar phrases, or as indicated elsewhere case) means a molecule that produces or enhances an immune response against a second molecule when it is connected to it. The term "analogous to" as used in the present context to define portions of a fusion protein, unless otherwise indicated, means "having the same or substantially the same structure as", for example, having the same or substantially the same amino acid sequence. For example, a protein portion that is analogous to a peptide endogenously synthesized by a vertebrate has the same or substantially the same amino acid sequence as the endogenously synthesized peptide.

"Substantially the same amino acid sequence" means a poly (amino acid) sequence that otherwise has the amino acid sequence of the peptide synthesized endogenously but in which one or more amino acid residues have been deleted, added or substituted by a different residue of amino acid, when the resulting poly (amino acid) molecule is useful for practicing the present invention. A poly (amino acid) molecule is useful for practicing the present invention if it can result in a specific immune response when it is in the product of the fusion protein. An amino acid substitution will preferably consist of conservative substitutions that are well known in the art. The rules for making such substitutions include those that have been described by Dayhof, M.D., 1978, Nat. Biomed. Res. Found., Washington, D.C., volume 5, supplement 3, among other citations. When a portion (a) or a portion (b) of a fusion protein of the present invention is cited in the present context as being "derived from" a peptide or pathogen, this means that the portion is analogous to the whole or to a part of the peptide or all or part of an immunogen (or antigen) from the pathogen, respectively. "A portion of" a peptide, antigen or immunogen for the purposes of this invention, unless otherwise indicated, is any part such that the resulting poly (amino acid) molecule is useful for practicing the present invention. This means that the part must be sufficient to produce an immune response against the pathogen from which it is derived (b) and / or against the peptide from which it is derived (a). The determination of these parts is within the ordinary experience in the technology sector. In a preferred embodiment, the peptide, antigen or immunogen portion constitutes at least 60%, more preferably 70%, and even more preferably at least 90% of the amino acid sequence of the particular peptide, antigen or immunogen. The actual percentage of the peptide, antigen or immunogen is less important than it is to include in the part the amino acid residues that will produce an immune response against (b) and / or (a). The terms "immunogen" and "antigen", as used in the present context, mean a molecule that is capable of eliciting an effective immune response in a vertebrate or a particular vertebrate species. The immunogens useful for the present invention are protein molecules, ie molecules that are composed of a sequence of amino acids, but which may also include non-protein groups, eg, carbohydrate residues. The term "immune response", for the purposes of this invention, means the production of antibodies and / or cells (such as T lymphocytes) that are targeted specifically or indirectly against, or that aid in the decomposition or inhibition of, a particular epitope. or several particular epitopes. An "effective immune response" is a immune response which, as far as portion (a) is concerned, is directed against one or more epitopes such that the activity of a peptide synthesized endogenously in the vaccinated vertebrate is inhibited; and as regards portion (b), it is directed against one or more epitopes of a pathogen in a manner that protects against the pathogen in the vaccinated vertebrate. "Triggering an immune response" and similar phrases, as used in the present context, mean to induce and / or enhance a immune response in a vertebrate in response to a vaccination. Phrases such as "inhibition of an infection" and "protection with respect to an infection" refer not only to the absolute prevention of an infection, but also to any detectable reduction in the degree or rate of infection by said pathogen., or any detectable reduction in the severity of the disease or any symptom or condition resulting from infection by the pathogen in the vaccinated animal as compared to an unvaccinated animal. A response that inhibits an infection can be induced in animals that have not been previously infected with the pathogen and / or that are not infected with the pathogen at the time of vaccination. It is intended that said phrases also include inhibiting the rate or degree of infection in an animal already infected with the pathogen at the time of vaccination. The term "double immune response" as used in the present context means an effective immune response as defined above, which inhibits the activity of more than one peptide, and preferably two different peptides, for example an endogenously synthesized hormone peptide. and a viral peptide. A "double-function vaccine", as used in the present context, means a vaccine that can produce a immune response in a vertebrate vaccinated therewith, which is directed against more than one peptide, and preferably against two different peptides, within of the vertebrate, for example a hormone synthesized endogenously by the vertebrate and a viral peptide of a virus that pathogenically infects the vertebrate. The phrase "endogenously synthesized peptide", as used in the present context and unless otherwise indicated, means a peptide that is synthesized by a vertebrate as part of the metabolic functioning of this vertebrate. Examples of endogenously synthesized peptides include, but are not limited to, hormones and enzymes. "Inhibiting the activity of a peptide" and similar phrases used herein, means interfering with the ability of the peptide to develop its normal function, for example its ability to catalyze a biochemical reaction (if the peptide is an enzyme), to trigger a biophysical response (if the peptide is a hormone), or to participate in infectivity or viral replication (if the peptide is a viral peptide). The phrases "amount effective to inhibit the activity of the peptide from which the portion (a)", "amount effective to inhibit the activity of GnRH", and the like, refer to the amount of a fusion protein. which is able to induce a immune response that is sufficient to interfere with the ability of the peptide to develop its function, such as preventing GnRH from stimulating or reducing the ability of GnRH to stimulate the release of LH or FSH, or to interfere with a surface protein of a virus so that it is unable to infect cells, thereby inhibiting replication and infection by the virus. An effective amount can be administered either as a single dose of a vaccine or as multiple doses of a vaccine. As used in the present context, the phrases "amount effective to inhibit an infection by the pathogen from which it is derived (b)", "amount effective to inhibit an infection by BHV-1", "effective amount to protect against an infection ", and other similar ones, refer to the amount of a fusion protein or a vaccine that is capable of protecting a vertebrate with respect to an infection as defined above. An effective amount can be administered either as a single dose of a vaccine or as multiple doses of a vaccine. A "vertebrate" as used in the present context, refers to any species that has a spine or spine, namely fish, amphibians, reptiles, birds and mammals. Examples of vertebrates that can benefit from the vaccine of the present invention, include, but are not limited to, humans, chickens, pigs, dogs, cats, cows, goats, lambs and horses, among others. Preferably, the vertebrate is a mammal. The term "pathogenically infecting" as used in the present context, refers to the ability of a pathogen to infect a vertebrate in a manner, or to such a degree, that it results in a morbid condition detectable in the vertebrate. BHV-1, for example, pathogenically infects animals of cattle, but not humans. Peptides that can be used as a source to prepare a portion (a) of a fusion protein of the present invention, include, but are not limited to, the following: 1) cholecystokines (Eng, J. et al., 1990, Regul. Pept. 30 (1): 15-9); a fusion protein of the present invention comprising a portion (a) analogous to all or a part of cholecystokines can be used to promote appetite in a vertebrate; 2) vasoactive intestinal peptide (Nilsson, A., 1975, FEBS Lett.60 (2): 322-6), whose inhibition causes a decrease in prolactin secretion which in turn discourages brooding behavior in chickens, resulting in an increased production of eggs; 3) a growth hormone and fragments of a growth hormone (Seeburg, P.H. et al., 1983, DNA 2 (1): 37-45); a fusion protein that enhances the activity of a growth hormone can promote growth in an animal; 4) a hormone that releases a growth hormone and its fragments (Gubler, U. et al., 1983, Proc.Nat.Acid.Sci.U.S.A. 80 (14), 4.311-4.314); the antibodies may also enhance the growth-promoting activity of a growth hormone-releasing hormone; 5) gastrin (Dimaline, R. et al., 1986, FEBS Lett 205 (2): 318-22; Kim, SJ et al., 1991, DNA Seq. 1 (3): 181 -7; Kariya, Y. collaborators, 1986, Gene 40 (1-3): 345-52) and a peptide that releases gastrin (Spindel, ER et al, 1986, Proc Nati Acad Sci USA 83 (1): 19-23); a fusion protein that inhibits the activity of gastrin and / or of a peptide that releases gastrin is useful, among other functions, to inhibit gastric secretions, and therefore to treat ulcers; treat cancers of the stomach, small intestine and / or colon; and to favor the appetite; 6) IgE peptides (Batista, F.D. et al., 1995, Nucleic Acids Res. 23 (23): 4.805-11); fusion proteins that inhibit IgE are useful for alleviating and / or preventing allergies, especially allergic skin reactions; 7) an angiotensin peptide, including angiotensin peptides I, II, III and IV (U.S. Patent 5,612,360 issued to Boyd et al., U.S. Patent 5,599,663 issued to Vaughan, U.S. Patent 5,629,292 issued to Rodgers and DiZerega, and U.S. Patent 5,635,359 issued to Brunner and Nussberger); a fusion protein that inhibits the activity of an angiotensin peptide is useful for treating, eg, hypertension in a mammal; 8) myostatin (Kambadur, R. et al., 1997, Genome Res. 7 (9): 910-6); inhibiting myostatin activity enhances the growth of the skeletal muscles in an animal, without impairing the quality of the meat, and therefore may be desirable to increase the production of meat in an animal; 9) inhibin or its fragments (U.S. Patent 5,786,179 issued to Kousoulas et al; U.S. Patent 5,665,568 issued to Mason and Seeburg); a fusion protein that inhibits the activity of inhibin can be used to treat infertility due to an irregular production of the follicle-stimulating hormone in a female animal; 10) somatostatin (U.S. Patent 5,422,110, cited above, the citations of Shen, LP et al., 1982, Proc. Nati. Acad. Sci. USA 79 (15): 4.575-9; Su, CJ et al., 1988, Mol Endocrinol 2 (3): 209-16); a fusion protein that inhibits somatostatin is useful, eg, to stimulate growth; and 11) cytokine peptides such as tumor necrosis factor (U.S. Patent 5,795,967 issued to Aggarwal et al.) and interleukin-1 (Masaaki, Y. et al., Japanese patent document JP 1994073095 -A 1 (filed on March 15, 1994)); inhibiting the activity of cytokines in an animal can alleviate immune-enhanced inflammation, for example an inflammation associated with allergies. The preceding peptides, their amino acid sequences and their physiological actions, are well known in technology. The aforementioned publications describing these peptides are incorporated herein by reference in their entirety. Examples of immunogens from which useful protein portions can be derived for portion (b) include, but are not limited to, the following immunogens: 1) OmpW (U.S. Provisional Patent Application No. 60 / 105,285, filed on October 22, 1998; encoded by the plasmid pER418 present in host cells of the Pz418 strain deposited in the American Type Culture Collection American Type Culture Collection (otherwise known as the ATCC (Manassas, Virginia, USA) under number 98928 and access to ATCC; SEQ ID NO: 44 (deduced amino acid sequence of OmpW)); OMPA1 (U.S. Provisional Patent Application No. 60 / 105,285, encoded by plasmid pER419 present in host cells of strain Pz419 deposited with the ATCC under accession number 98929 to the ATCC, SEQ ID NO: 45 (deduced amino acid sequence of OmpA1)); OmpA2 (U.S. Provisional Patent Application No. 60 / 105,285), encoded by plasmid pER420 present in host cells of strain Pz420 deposited with the ATCC under the designation number 98930 of access to the ATCC, SEQ ID NO. : 46 (deduced amino acid sequence of OmpA2)); OmIA serotype 1 and serotype 5 (U.S. Patent No. 5,441,736 issued to Gerlach et al.); all from Actinobacillus pleuropneumoniae; a protein portion analogous to all or a portion of the OmpW, OmlA5 or OmpA can be used as a support in a fusion protein according to the present invention while simultaneously providing the swine with protection against porcine pleuropneumonia (caused by a infection by A. pleuropneumoniae); 2) hepatitis B surface antigen (Hsiung et al., 1984, J. Mol. Appl. Gen. 2: 497); a protein portion analogous to all or a portion of a hepatitis B surface antigen can be used as a support in a fusion protein of the present invention while simultaneously providing protection in humans against a hepatitis B infection; 3) an RTX toxin ("repeat in toxin") from Actinobacillus pleuropneumoniae (Frey, J. et al, 1991, Infect. Immun 59 (9), 3026-32); a protein portion analogous to all or a portion of an RTX toxin as a support in a fusion protein of the present invention can simultaneously provide immune protection against Actinobacillus pleuropneumoniae in swine and animals of cattle; 4) the subunit of labile enterotoxin against the heat of E. coli (Leong, J. et al, 1985, Infect Immun 38 (1): 73-7, Inoue, T. et al., 1993, FEMS Microbiol. Lett 108 (2): 157-61); a portion analogous to all or a portion of the subunit of labile enterotoxin against the heat of E. coli may serve as a support that also provides immune protection against E. coli in swine and cattle animals; 5) E. coli K88 pilus or K99 pilus antigens (Bakker, D. et al., 1992, J. Bacteriol 174 (29): 6,350-8; Simons, BL et al., 1990, FEMS Microbiol. Lett 55 (102 ): 107-12); a protein portion analogous to all or a portion of the K88 pilus antigen or the K99 pilus antigen as a support in a fusion protein of this invention may provide protection against an enteric disease caused by E. coli in swine and cattle animals bovine; 6) the p68 antigen of B. bronchiseptica (WO 9115571 -A 5 (October 17, 1991)); a protein portion analogous to all or a portion of the p68 antigen may be used as a support in a fusion protein of the present invention and may provide protection against borderline infection (kennel cough disease) in canine animals; 7) glycoprotein 53 from bovine viral diarrhea virus (BVD of Bovine Viral Diarrhea) (Fritzemeier, J. et al, 1997, Arch. Virol. 142 (7): 1335-50); a portion analogous to all or a portion of the glycoprotein 53 can serve as a support in a fusion protein and also provide protection from fatal mucosal disease in cattle animals; 8) viral proteins 1 and 2 of parvovirus (Xie, A and Chapman, M.S., 1996, J. Mol. Biol. 264: 497); a protein portion analogous to all or a portion of the viral protein 1 or of the viral protein 2 relative to parvovirus can serve as a support in a fusion protein of the present invention and simultaneously protect suids, dogs and cats with respect to of a parvovirus infection; a coronavirus binding protein (Kokubu, T. et al, 1998, Journal of the Japan Veterinary Medical Association 51: 252-55; Lewis, EL, 1996, Thesis presented at the University of Bristol (University of Bristol (Clifton, Bristol , United Kingdom = UK)), Britton, P. et al., 1991, Virus Res, 21 (3): 181-98); a portion analogous to all or a portion of the coronavirus binding protein can be used as a support in a fusion protein and also provides protection against a coronavirus infection in animals of cattle, suids, dogs and cats; 10) an outer membrane protein regulated by bacterial iron (Gerlach, GF et al., 1992, Infect.Immunol.60 (8): 3.253-61; Thompson, SA et al., 1993, Mol. Microbiol. 9 (1): 85-96); a portion analogous to all or a portion of said membrane protein can be used as a support which also provides immune protection against Actinobacillus pleuropneumoniae and / or meningitis in swine, cattle and poultry animals; 11) the rabies G protein (Shinichi, S. et al., JP 198917489-A 1 (filed July 6, 1989)); a protein portion analogous to all or a portion of the rabies G protein can be used as a support in a fusion protein and will also simultaneously provide protection in cats, dogs and wild animals with respect to rabies; 12) plasminogen activator protein from Streptococcus uberis (Leigh, J.A., 1993, WO 9314209); a protein portion analogous to all or a portion of the plasminogen activator protein of Streptococcus uberis is useful as a support and will also provide treatment and / or protection with respect to mastitis in dairy cows; 13) the influenza virus hemagglutinin protein (Hovanec, DL and Air, GM, 1984, Vírology 139 (2): 384-92) and the nucleocapsid protein of the influenza virus (Lindstrom, SE et al., 1998, J. Virol. 72 (10): 8.021-31); a portion analogous to all or a portion of any of these proteins can be used as a support in a fusion protein of this invention and simultaneously provide immune protection against influenza in humans, suids and poultry; 14) tetanus toxoid (Fairweather, N. F. et al., 1986, J. Bacteriol 165 (1): 21-7, Niemann, H., 1986, EMBO J. 5 (10): 2495-502); a protein portion analogous to all or a portion of the tetanus toxoid can be used as a support in a fusion protein that will also provide protection in humans, horses and animals of cattle against tetanus; 15) pertussis toxoid (Nicosia, A. et al., 1986, Proc. Nati, Acad. Sci. USA 83 (13): 4.631-5); a protein portion analogous to all or a portion of the pertussis toxoid can serve as a support in a fusion protein and will provide immunity protection with respect to pertussis in humans; 16) a herpes virus glycoprotein (Gompels, UA et al., 1992, DNA Seq. 3 (1): 25-39; Misra, V. et al., 1988, Virology 166: 542-9; Whitbeck, JC, and collaborators, 1988, J. Virol., 62: 3319-27; Fítzpatrick, DR et al., 1989, Virology 173: 46-57); a protein portion analogous to all or a portion of a herpes virus glycoprotein may serve as a support in a fusion protein of this invention and may also function in the fusion protein to provide immune protection with respect to herpes in humans and cattle animals; 17) the intimin protein of enterohemorrhagic E. coli (Jerse, A.E. et al., 1990, Proc. Nati, Acad. Sci. USA 87 (20): 7,839-43); a portion analogous to all or a portion of the intimin protein of enterohemorrhagic E. coli may function as a support and also provide protection with respect to a hemorrhagic disease in species that include humans and animals of cattle; 18) VP2 (Cao, Y.C. et al., 1995, Ping Tu Hsueh Pao 11 (3): 234-41); a portion analogous to all or a portion of VP2 can function as a support and can also provide immune protection with respect to infectious bursa disease in poultry; and 19) the F and G proteins of the respiratory syncytial virus (Schrijver, RS et al., 1997, Archives of Virology 142 (11): 2195-2210; Furze, JM et al., 1997, Virology 231 (1): 48- 58); a protein portion analogous to all or a portion of the F protein or the G protein can act as a support and will also provide immunity protection with respect to the Bovine Respiratory Syncytial Virus in cattle animals. The foregoing immunogens and their amino acid sequences are known in the field of technology. The aforementioned publications that describe the immunogenic precedents are incorporated herein by reference in their entirety. Different protein portions (a) and (b), each portion being analogous to all or a portion of a peptide or immunogen described in one of the preceding paragraphs or another known peptide or immunogen, may be combined according to the present invention. to form a fusion protein specifically designated for a particular vertebrate, eg, a cow, a pig, a chicken or a human, or a particular category of vertebrates, eg, mammals or primates, to inhibit activity of a particular peptide in the vertebrate while simultaneously protecting the vertebrate with respect to an infection by a certain pathogen. As an example, GnRH is a hormone of the reproductive system synthesized by cattle. Inhibition of GnRH activity in cattle animals will provide a beneficial reduction in the expression of sexual characteristics such as aggressive behavior. Since BHV-1 pathogenically infects cattle animals, an immunogen from BHV-1 can be used as a carrier with GnRH. Therefore, in one embodiment, a portion (a) analogous to all or a portion of a GnRH peptide and a portion (b) analogous to all or a portion of an immunogen from BHV-1 are connected to provide a fusion protein that induces a double immune response in cattle animals that inhibits the activity of GnRH and also protects against an infection with BHV-1. In another non-limiting example, the present invention provides a fusion protein in which the portion (a) is analogous to all or a portion of a growth hormone, and in which the portion (b) is analogous to the whole or to a part of a BHV-1 antigen. Said fusion protein is useful for regulating growth in cattle animals while providing a protective immune response with respect to BHV-1. In another example, portion (a) is analogous to all or part of the IgE peptide and portion (b) is analogous to all or a portion of the p68 antigen of B. bronchiseptica. The resulting fusion protein is useful for treating or preventing allergies, especially allergic skin reactions, in dogs, while providing a protective immune response with respect to bordetella. In yet another example, portion (a) is analogous to all or a portion of cholecystokines and portion (b) is analogous to all or a portion of OmpW, OmIA serotype 1, OmIA serotype 5, OmpA1 or OmpA2 from Actinobacillus pleuropneumoniae. Said fusion protein is useful to excite the appetite in swine while simultaneously providing a protective immune response with respect to porcine pleuropneumonia. The protein portions (a) and (b) for the fusion proteins of the invention can be obtained according to methods known in the field of technology. For example, either or both of portion (a) or portion (b) can be obtained by purification from natural sources. Alternatively, either or both of portion (a) or portion (b) can be obtained by synthetically linking amino acids together. Alternatively, either or both of portion (a) or portion (b) can be recombinantly synthesized using well-known recombinant techniques from a polynucleotide molecule comprising a nucleotide sequence that encodes the portion (a) ) or portion (b). Preferably, a polynucleotide molecule comprising a nucleotide sequence encoding portion (a) is linked to a polynucleotide molecule comprising a nucleotide sequence that encodes portion (b), such that the entire fusion protein is synthesized recombinantly. Recombination techniques within ordinary experience in the technology sector can be used to prepare polynucleotide molecules that encode portions (a) and (b) of the fusion proteins in question. These techniques are described, among other sites, in the citations of Maniatis, et al., 1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Ausubel, et al., 1989, Current Protocols in Molecular Biology, Greene Publishing Associates & Wiley Interscience, NY; Sambrook, et al., 1989, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY; Innis and collaborators (editing coordinators), 1995, PCR Strategies, Academic Press, Inc., San Diego; and Erlich (editing coordinator), 1992, PCR Technology, Oxford University Press, New York, all of which are incorporated herein by reference. The amino acid sequences of many hormonal peptides are well known in the field of technology. Some known hormonal peptides are described above. As another example, the amino acid sequence of GnRH is known in the field of technology (see, eg, Ladd, A., 1993, cited above). The amino acid sequence of GnRH is also provided in the present text (SEQ ID NO: 13). Alternatively, if the amino acid sequence of a hormone is not known, it can be determined using classical techniques, such as performing repeated cycles of Edman degradation in a purified protein fraction followed by an amino acid analysis using an HPLC (high pressure liquid). chromatography - high pressure liquid chromatography) (see, e.g., U.S. Patent 5,422,110, supra). Similarly, a protein portion that is the same or substantially the same as an immunogen from a pathogen can be obtained according to classical techniques, from a known amino acid sequence or by determining the amino acid sequence as described above. A protein portion that is substantially the same as an immunogen from a pathogen can be determined, for example, by comparing the amino acid content of the protein portion of the known amino acid content of the immunogen, or by performing a sequence alignment that compares the protein portion with the amino acid sequence of the immunogen, using known techniques. Examples of BHV-1 antigens, from which the protein portion (b) can be derived, include, but are not limited to, the gB of BHV-1, gC of BHV-1 and gD of BHV-1 ( also known in the art sector as gl, glll and glV of BHV-1, respectively). Methods for obtaining protein portions that are analogous to all or a portion of said antigens have been described above. For example, U.S. Pat. 5,151,267 issued to Babiuk et al. Describes the nucleotide sequences and deduced amino acid sequences of the gl, glll and glV of BHV-1. See also U.S. Pat. 5,585,264 issued to Babiuk et al. In addition, U.S. Pat. 5,545,523 issued to Batt et al. Describes specific oligonucleotides for BHV-1 that are useful in the amplification of the gene sequences of the gl and glV of BHV-1. In addition, methods for purifying BHV-1 glycoproteins from virus-infected cell cultures have been described (Babiuk, L.A. et al., 1987, Virology 159: 57-66). The amino acid sequence of the gD of BHV-1 full length as published in the citation of Tikoo et al., 1990, supra, is provided herein (see Figure 5 and SEQ ID NO: 19). The expression of the mature gD of full-length BHV-1 has been carried out in baculovirus, adenovirus, vaccinia virus and E. coli systems (van Drunen Littel-van den Hurk, S. et al., 1993, Vaccine 1 1: 25-35). The disclosures and teachings of the aforementioned patents and publications are incorporated herein by reference. Another example of a gD antigen of BHV-1 that is useful, in whole or in part, for a fusion protein of the present invention is the poly (amino acid) of gD of BHV-1 encoded by the clone FlgD / Pots207nco (# 79) (see Figure 3 and SEQ ID NO: 17). Although any part of a BHV-1 antigen that is capable of stimulating an immune response that inhibits the peptide from which portion (a) is derived and, as in the first aspect of the invention, an immune response that protects a cow with respect to a BHV-1 infection can be used in the fusion proteins of this invention, examples of preferred parts of the gD of BHV-1. which can be used in this invention are truncated gD (tgD), mature gD (mgD) and truncated mature gD (tmgD). Truncated gD (tgD) refers to a gD protein in which the transmembrane domain, optionally with nucleotides located downstream and / or upstream, has been completely or partially deleted. The transmembrane domain for gD is known in the field of technology as a region of a particular poly (amino acid) of a gD of generally very hydrophobic amino acids. The transmembrane domain for gD / Pots, the gD of BHV-1 encoded by the clone FlgD / Pots207nco (# 79), is reproduced in Figure 3. The transmembrane domain for gD / Pots starts at amino acid 364 (valine) and ends in amino acid 389 (tyrosine). The term "mature gD" refers to a gD protein that has no signal sequence at the amino terminus. The signal sequence of full length gD / Pots is reproduced in Figure 3. In another embodiment, the protein portion (b) of the fusion protein of the present invention may comprise a heterologous signal sequence attached to the terminal end of amino of the protein. Alternatively, portion (b) may not comprise any signal sequence. In one embodiment of the invention, portion (b) is analogous to a gD of BHV-1 that is both truncated and mature (tmgD). An example of a truncated mature gD antigen is provided in SEQ ID NO: 35. An example of a truncated gD antigen that is not mature is provided in SEQ ID NO: 29. As used in the present context, " tgD "refers to a gD protein of BHV-1 that is truncated as described above," mgD "refers to a gD protein of BHV-1 that is mature as described above, and" tmgD "refers to a gD protein of BHV-1 that is both truncated and mature. The term "GnRH peptide" means, unless otherwise indicated, a molecule having an amino acid sequence of SEQ ID NO: 13. In one embodiment, the fusion proteins in question comprise multiple portions (a) analogous to a GnRH peptide. In preferred embodiments, the fusion proteins of this invention comprise one or more portions analogous to four GnRH peptides sequentially linked, ie, one or more GnRH-tetramer-analogous portions. In a preferred embodiment, a fusion protein of the present invention comprises a portion of 4GnRH. As used in the present context, "4GnRH" refers to a GnRH tetramer having four GnRH peptides sequentially linked in the same amino terminal / carboxy terminal end orientation. Preferably, the fusion proteins of the present invention comprise one or more tetramers of GnRH, each tetramer having the amino acid sequence shown in SEQ ID NO: 15. The expressions provided with hyphens provided herein and containing the terms "4GnRH", " tmgD "," tgD "and" mgD "(as defined above) indicate fusion proteins comprising portions of a poly (amino acid) corresponding to the terms linked from left to right in the order indicated, the left end corresponding to the extreme amino terminal of the fusion protein and the right extremity corresponding to the carboxy terminus of the fusion protein. The portions of a poly (amino acid) can be linked directly to each other or can be linked indirectly, ie the portions can be separated by one or more (for example from 1 to 10, preferably from 1 to 3) amino acids. Therefore, "tmgD-4GnRH" refers to a fusion protein having a truncated mature gD portion connected, directly or indirectly, with a 4GnRH portion, the carboxy terminus of the truncated mature gD portion being linked (directly or indirectly) to the amino terminal end of 4GnRH. As another example, "tgD-4GnRH" refers to a fusion protein having the amino terminus of the 4GnRH portion connected to the carboxy terminus of a truncated gD antigen that is not mature. As another example "4GnRH-tmgD-4GnRH" refers to a portion of 4GnRH having a carboxyl terminus bonded to the amino terminus of a portion of tmgD, whose portion of tmgD in turn is linked by its carboxyl terminus to the terminus of amino of a second portion of 4GnRH. The fusion proteins of the present invention include, but are not limited to, the examples of fusion proteins described in this paragraph. Another example of a fusion protein of this invention is tmgD-4GnRH. In any of the aforementioned examples, the portions may be linked directly or indirectly. As discussed above, protein portions (a) and (b) can be chemically connected through chemical linkers and techniques that are well known in the technology sector. As an example, certain amino acids existing in a portion (a) or (b), for example in a portion (b) analogous to gD, can be activated with a reagent, such as iodoacetamide. The remaining portions (a) or (b), for example monomers and / or multimers of GnRH, can be added. In this example, the cysteine residues terminally incorporated in the GnRH react with activated lysine residues in the gD-like protein. This reaction results in fusion proteins according to the present invention comprising a portion analogous to central gD having multiple GnRH analogues connected around it in various lysine residues. In another example, portions (b) analogous to a BHV-1 antigen can be combined together with portions (a) analogous to monomers or multimers of GnRH in the presence of ethyl-dimethylaminopropyl-carbodiimide (EDAC) and N-hydroxy -succinimide (NHS) (see Bernatowics, M. and Matsueda, G., 1986, Analytical Biochemistry 155: 95-102). This reaction also results in a central portion (b) analogous to all or a part of an antigen of BHV-1 with multiple portions (a) analogous to monomers or multimers of GnRH chemically connected to it. The chemically synthesized fusion proteins of the present invention can also optionally be chemically modified to comprise substituents other than amino acids, for example carbohydrate substituents using known techniques. Other chemical techniques for combining protein portions, either with multiple linkages to a protein center or linear linkages of protein portions, can be used to chemically synthesize fusion proteins of the present invention using known techniques. Techniques for preparing chemically synthesized fusion proteins of the present invention are described, inter alia, in Dunn and Pennington, 1994, Methods in Molecular Biology, volume 26, chapter 10 (Humana Press Inc.), which is incorporated herein by reference. the present for your reference. The present invention also comprises recombinant fusion proteins as described above. Examples of recombinant fusion proteins according to the present invention include the recombinant fusion protein encoded by the plasmid pCMV-gD: GnRH, the plasmid pQE-gD: GnRH, the recombinant fusion protein encoded by the plasmid pQE-GnRH: gD : GnRH and the recombinant fusion protein encoded by the plasmid pQE-GnRH: gD. The cells containing these plasmids have been deposited in the American Type Culture Collection (ATCC Manassas, Virginia, USA); they have been assigned the access numbers 203370, 98953, 98955 and 98954, respectively. Another example of a recombinant fusion protein of the present invention is the recombinant fusion protein expressed by the baculovirus artificial structure Bac-gD: GnRH. The Bac-gD: GnRH has also been deposited with the ATCC and has been assigned the number VR-2633 to access the ATCC. The aforementioned pQE plasmids and the artificial baculovirus structure are particularly useful for the in vitro expression of fusion proteins. Plasmid pCMV-gD: GnRH is particularly useful for expression in vivo. The recombinant fusion proteins according to this invention may optionally comprise portions that help to purify the fusion proteins from the reaction medium following transcription and in vitro translation. An example of a poly (amino acid) sequence that can aid in the purification of a recombinant protein from the medium is a 6XHIS tag. The phrase "6XHIS mark" is used in this application interchangeably with "6XHIS leader". The sequence of the 6XHIS tag encoded by the pQE-31 vector is provided in SEQ ID NO: 37. The proteins comprising a 6XHIS tag can be purified from the media by passing these media through a nickel column such as a Ni-NTA column from Qiagen (Chatsworth, CA). Another example of a portion that can help purify recombinant fusion proteins of this invention as a consequence of in vitro expression is the FLAG® epitope tag (International Biotechnologies, Inc., New Haven, CT) which is a hydrophilic label peptide. . The gene encoding the FLAG® epitope tag can be introduced by classical techniques into a polynucleotide molecule comprising a nucleotide sequence encoding a fusion protein of this invention at a corresponding point, eg, to the terminal end of amino or carboxyl of the fusion protein. A fusion protein expressed therefrom can then be detected and purified by affinity using commercially available anti-FLAG® antibodies. Other means for purifying recombinant proteins expressed in vitro are well known in the art and can be used to purify the recombinant fusion proteins of the present invention. These methods are described, inter alia, in those of Marshak, D.R. et al., 1996, Strategies for Protein Purification and Characterization: a Laboratory Course Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY. Once a fusion protein of the present invention has been obtained, it can be characterized, if desired, by classical methods, including by SDS-PAGE, size exclusion chromatography, analysis of amino acid sequences, etc. The fusion protein can be further characterized using a hydrophilicity analysis (see, eg, Hopp and Woods, 1981, Proc. Nati, Acad. Sci, USA 78: 3824), or algorithms of analog software software programs, to identify hydrophobic and hydrophilic regions. A structural analysis can be carried out to identify regions of the fusion protein that adopt specific secondary structures. Biophysical methods such as X-ray crystallography (Engstrom, 1974, Biochem Exp. Biol. 1 1: 7-13), computer modeling (Fletterick and Zoller (editing coordinators), 1986 in: Current Communications in Molecular Biology, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY) and nuclear magnetic resonance (NMR) can be used to map and study potential sites of interaction between the polypeptide and other proteins / receptors / putative interacting molecules such as antibodies.

Polynucleotide Molecules and Vectors Encoding Fusion Proteins The present invention further provides a polynucleotide molecule comprising a nucleotide sequence encoding a fusion protein of the present invention. Examples of polynucleotide molecules of this type include, but are not limited to, a polynucleotide molecule comprising the nucleotide sequence set forth in SEQ ID NO: 34, which encodes a fusion protein 4GnRH-tmgD; a polynucleotide molecule comprising the nucleotide sequence set forth in SEQ ID NO: 40, which encodes a fusion protein 4GnRH-tmgD-4GnRH; and a polynucleotide molecule comprising the nucleotide sequence set forth in SEQ ID NO: 41, which encodes a tmgD-4GnRH fusion protein. The present invention also provides cloning and expression vectors comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding a fusion protein of the invention. The term "vector", as used in the present context, means a unit comprising genetic information (in the form of polynucleotide sequences), whose information is capable of expressing a poly (amino acid) and / or programming the replication of the unit when appropriate conditions and resources are present (eg, amino acids, nucleotides, and transcription factors). Examples of such units include viruses, plasmids and cosmids. As used in the present context, the terms "nucleotide sequence", "coding sequence", "polynucleotide", "polynucleotide sequence", and the like, refer to both DNA and RNA sequences, which can be either either single-stranded or double-stranded, and may include one or more prokaryotic sequences, eukaryotic sequences, cDNA sequences, genomic DNA sequences, including exons and introns, and chemically synthesized DNA and RNA sequences. The production and manipulation of polynucleotide molecules of the present invention comprising nucleotide sequences encoding portions (a) and (b) of the fusion proteins in question are within the ordinary experience in the field of technology and they can be carried out according to recombinant techniques described, inter alia, in those of Maniatis et al., cited above; Ausubel et al., Cited Sambrook et al., Cited above; Innis et al., Cited above; and Erlich, cited above. The nucleotide sequences encoding many peptides of hormones and peptides of viral antigens are known in the field of technology, and such information can be used to prepare coding regions for protein portions (a) and (b). Said sequences are presented, inter alia, in the aforementioned references that describe immunogens and peptides useful in the present invention. Alternatively, the nucleotide sequences of peptides and viral antigens can be deduced using methods known in molecular biology. The nucleotide sequences encoding portion (a) and / or portion (b) can be prepared by synthesis. The desired sequence can be prepared from overlapping oligonucleotides. See, e.g., Edge, 1981, Nature 292: 756; Nambair et al., 1984 Science 223: 1299; Jay et al., 1984, J. Biol. Chem. 259: 6,311 and U.S. Pat. 5,422,110, cited above. As another example, the amino acid sequence of a peptide or antigen can be used to design probes intended to identify the gene encoding the peptide or antigen in a genomic library. In this method, oligonucleotide probes encoding a portion of the amino acid sequence of the peptide or antigen are prepared. Oligonucleotide probes are used to screen an appropriate DNA library for genes encoding the peptide or antigen. Generally, the DNA library that is screened is a library prepared from a genomic DNA or a genomic RNA (cDNA) from an appropriate source, such as from a cell or tissue expressing or from a virus encoding the antigen. The techniques for isolating genes in this way are well known in the field of technology. Nucleotide sequences that are homologous with sequences obtained as described above to encode immunogens or peptides can also be used in the present invention. For the purposes of the present invention, a second nucleotide sequence is "homologous" with a first nucleotide sequence when it encodes the same protein, the same peptide or other poly (amino acid) other than the first nucleotide sequence, or when it encodes a nucleotide sequence. poly (amino acid) that is sufficiently similar to the poly (amino acid) encoded by the first nucleotide sequence to be useful in the practice of the present invention. Since the genetic code is degenerate, a homologous sequence of nucleotides can include any number of "tacit" base changes, ie, nucleotide substitutions that nevertheless encode the same poly (amino acid). A homologous nucleotide sequence may also contain non-tacit mutations, ie substitutions, deletions or additions of bases resulting in amino acid differences in the poly- (amino acid) encoded, as long as the poly (amino acid) sequence remains useful for practicing the present invention. A second nucleotide sequence that is homologous with a first nucleotide sequence is preferably one that hybridizes with the complement of the first nucleotide sequence under moderately stringent conditions, i.e., by hybridization to a DNA attached to a filter in NaHP04. , 5 M, sodium dodecyl sulfate (SDS, sodium dodecyl sulfate) 7%, EDTA 1 mM at 65 C, and washing in 0.2xSSC / 0.1% SDS at 42 C (see Ausubel et al., previous appointment). More preferably, homologous sequences of nucleotides hybridized to each other under very stringent conditions, i.e., by hybridization to a DNA fixed to a filter in 0.5 M NaHP04, 7% SDS, 1 mM EDTA at 65 C, and washing in 0.1xSSC / 0.1% SDS at 68 C (Ausubel et al., Cited above). After having obtained polynucleotide molecules comprising nucleotide sequences encoding portions (a) and (b), these polynucleotide molecules can be ligated together using appropriate enzymes and techniques known to form a polynucleotide molecule comprising a nucleotide sequence which encodes a fusion protein of this invention. Examples of coding sequences useful for constructing polynucleotide molecules comprising sequences encoding fusion proteins of the present invention, and vectors comprising said polynucleotide molecules, include, but are not limited to, the sequence presented in SEQ ID NO: 16 , which encodes the FlGD / Pots antigen of gD of BHV-1, expressed by clone FlgD / Pots207nco (# 79), set forth in SEQ ID NO: 17; the sequence presented in SEQ ID NO: 18, which encodes the gD of BHV-1 M59846, set forth in SEQ ID NO: 19; the sequence presented in SEQ ID NO: 28, which encodes a truncated gD antigen, which is not mature, set forth in SEQ ID NO: 29; and the sequence presented in SEQ ID NO: 36 encoding a truncated mature gD, set forth in SEQ ID NO: 35. An example of a nucleotide sequence encoding a GnRH monomer is set forth in SEQ ID NO: 33. An example of a sequence encoding a GnRH tetramer, namely the GnRH tetramer having the amino acid sequence set forth in SEQ ID NO: 15, is set forth in SEQ ID NO: 32. In one embodiment, a vector of the present invention is suitable for the in vitro expression of a fusion protein, such as a plasmid that is capable of transfecting a host cell such as a bacterial cell and expressing the fusion protein in the bacterial cell. Examples of plasmid vectors include plasmids such as recombinant pQE plasmids, capable of transfecting bacteria and expressing the fusion proteins of this invention. Examples of some prokaryotic expression vector plasmids into which a polynucleotide molecule comprising a nucleotide sequence encoding a fusion protein of the invention can be introduced, include pQE-50 and pQE-31 (Qiagen, Chatsworth, CA), pUC8, pUC9, pBR322 and pBR239 (Biorad Laboratories, Richmond, CA), pPL and pKK223 (Pharmacia Piscataway, NJ). Other plasmids known in the art can also be used to prepare vectors comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding a fusion protein of this invention, and said plasmids can be determined by which They have ordinary experience. Preferred plasmids which are capable of expressing fusion proteins of the invention in vitro include pQE-gD: GnRH (accession number 98953 to the ATCC), pQE-GnRH: gD: GnRH (accession number 98955 to the ATCC), and pQE -GnRH: gD (access number 98954 to the ATCC). These plasmids are capable of expressing fusion proteins of this invention in E. coli bacteria. In another embodiment, a vector of the present invention is a plasmid suitable for in vivo expression of a fusion protein. Plasmids that are capable of transfecting eukaryotic cells, and that can be used to construct vectors of the present invention, can be determined by those with ordinary experience in the field of technology. Said plasmids may comprise sequences and encode elements that aid in the expression and in vivo treatment of the fusion proteins in a vaccinated vertebrate. For example, a plasmid of the present invention may comprise a eukaryotic promoter sequence. As another example, a plasmid of the present invention may comprise a sequence encoding a signal bound to the expressed fusion protein, whose signal results in the transport of the expressed fusion protein to the cell membrane and the excretion of the protein from fusion from the cell to the circulatory system of the vaccinated vertebrate. An example of a plasmid that can be used to construct vectors of the present invention, capable of expressing fusion proteins in vivo, is pCMV (Clontech, Inc., Palo Alto, CA). Other typical eukaryotic expression plasmids that can be engineered to comprise a polynucleotide molecule comprising in turn a nucleotide sequence encoding a fusion protein of the present invention, include an inducible expression system in a mammal and systems based on cytomegalovirus promoter-enhancer (Promega, Madison, WE; Stratagene, La Jolla, CA; Invitrogen). Other plasmids useful for preparing vectors expressing in vivo fusion proteins of the present invention can be determined by those having ordinary experience in the field of technology. A preferred example of a plasmid of the present invention, capable of expressing a fusion protein in vivo is pCMV-gD: GnRH that has been deposited with the ATCC (accession number 203370 to the ATCC). The vectors of the present invention can also include recombinant viruses comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding a fusion protein of the present invention. Said viruses can be prepared according to techniques known in the field of technology. They can be prepared for example from a bacteriophage, the resulting recombinant bacteriophage being useful for expressing and producing in vitro the fusion proteins in question within bacteria. Examples of bacteriophages that can be used to prepare vectors of this invention include Ta, T7, X174, G4, M13 and fd. Other bacteriophages useful for the present invention can be determined by those with ordinary experience in the technology sector. Recombinant viruses capable of transfecting insect cells or yeast cells can also be constructed for the expression and in vitro production of the fusion proteins of this invention in insect cells and yeast cells, respectively. In this regard, another example of a vector that can be used for the in vitro production of the fusion proteins of this invention is a recombinant virus based on a baculovirus. In preferred embodiments, the present invention provides baculovirus vectors that express tmgD-4GnRH, 4GnRH-tmgD-4GnRH or 4GnRH-tmgD. In a preferred embodiment of this invention, the vector is the Bac-gD: GnRH baculovirus vector, which expresses a tmgD-4GnRH fusion protein. The Bac-gD: GnRH has been deposited with the ATCC (number VR-2633 to access the ATCC). Recombinant viruses capable of infecting and expressing the fusion proteins in question in eukaryotic cells, such as avian or mammalian cells, including viruses for both in vitro and in vivo expression of fusion proteins within eukaryotic cells, they can also be built according to techniques well known in the technology sector. Examples of viruses from which such recombinant viruses can be prepared include poxviruses, such as vaccinia virus, and adenoviruses. Both recombinant vaccinia viruses and recombinant adenoviruses can be used for expression either in vitro or in vivo. Other viruses suitable for expression in eukaryotic cells can be determined by those with ordinary experience in the field of technology. In another embodiment, a vector of the present invention is a "transfer vector" comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding a fusion protein of the present invention. A transfer vector is a plasmid comprising a sequence encoding a peptide, the plasmid of which can infect an appropriate host cell, such as an appropriate insect or mammalian cell, in a process of co-infection in vitro with a virus, which gives place where the host cell produces a recombinant virus, whose recombinant virus is itself a vector that is capable of expressing the peptide encoded by the plasmid in an appropriate expression system. The preparation of transfer vectors for the in vitro production of recombinant viruses is well known in the field of technology, and the plasmids that are useful for preparing transfer vectors according to this invention in question can be determined by those with experience. ordinary in the technology sector. Examples of suitable plasmids for preparing transfer vectors include, but are not limited to, pBacPAKβ and pBacPAK9 (Clontech, Inc.). A preferred transfer vector for preparing a viral vector encoding a fusion protein of the present invention is the transfer vector pBacHISgD: GnRH. The nucleotide sequence encoding a fusion protein of the present invention can be linked to, and placed under the control of, various nucleotide elements, such as signal sequences, inducible and non-inducible promoters, ribosome binding sites for bacterial expression, and operators. Said elements allow the nucleotide sequence to be transcribed either in vivo or in vitro, in a host cell transfected with a vector comprising the polynucleotide molecule, and correspondingly cloned or expressed in the host cell. Regulatory sequences and enhancer sequences can also be included in the polynucleotide molecules of the invention. The coding sequences are put into "operational association" with the elements that are included in the polynucleotide molecules, which means that their positioning and orientation are such that a transcription of the coding sequences can occur. This placement is within the ordinary experience in the technology sector. The regulatory elements of polynucleotide molecules of the present invention can vary in their strength and their specificities. Depending on the host system and vector to be used, any of a number of appropriate transcription and translation elements can be used. For example, when cloning into mammalian cell systems, promoters isolated from the genome of mammalian cells, eg, the mouse metallothionein promoter, or from viruses growing on these cells, can be used, the 7.5 K promoter of the vaccinia virus or the long terminal repeat of Moloney murine sarcoma virus. Promoters obtained by recombinant DNA or synthesis techniques can also be used to provide transcription of the introduced sequence. In addition, expression from certain promoters can be elevated in the presence of particular inducers, eg, zinc and cadmium ions for metallothionein promoters. Non-limiting examples of regions or promoters that regulate transcription include, for bacteria, the -gal promoter, the T7 promoter, the T5 promoter, the TAC promoter, the left and right promoters, the trp and lac promoters , the fusion promoters of trp-lac, etc; for yeasts, glycolytic enzyme promoters, such as the ADH-I and -II promoters, the GPK promoter, the PG1 promoter, the TRP promoter, etc .; and for mammalian cells, the SV40 early and late promoters, the major adenovirus late promoters, among others. Specific initiation signals can also be used for the translation of introduced coding sequences. These signals typically include an ATG start codon and adjacent sequences. In cases where the polynucleotide molecule of the present invention includes its own initiation codon and adjacent sequences are introduced into the appropriate expression vector, no additional translation control signal may be required. However, in cases where only a portion of a coding sequence is introduced, exogenous translation control signals, including the ATG start codon, may be required. These exogenous signals of translation control and these initiation codons can be obtained from a variety of sources, both natural and synthetic. In addition, the initiation codon must be in phase with the reading frame of the coding regions to ensure an in-frame translation of the entire insert. The vectors of this invention may also include genes and repressor operators, which regulate the transcription of an mRNA. Examples of operators that can be included in the vectors in question include the sequence of the lac operator. Other operators are known in the field of technology, and these can be included within the vectors of this invention. The expression vectors may also contain a polynucleotide molecule of this invention that is further engineered to contain polylinker sequences that encode specific protease cleavage sites such that the expressed fusion protein can be released from sequences of vectors expressed by treatment with a specific protease. For example, the fusion protein vector can include a nucleotide sequence encoding a thrombin or a factor Xa cleavage site, among others. The expression vectors of the present invention can also comprise nucleotide sequences that encode a poly (amino acid) that can help purify a fusion protein from media as a continuation of an expression. An example of such a nucleotide sequence is a nucleotide sequence encoding a 6XHIS tag, such as the nucleotide sequence set forth in SEQ ID NO: 38.

Transformed Cells for Expressing Fusion Proteins The present invention also provides transformed cells comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding a fusion protein as described herein. Cells useful for transformation by this invention include bacterial cells, yeast cells, mammalian cells, insect cells and plant cells. The transformed cells of this invention can be prepared by transfecting a cell with a vector comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding the fusion protein as described above. Host cells useful for practicing the present invention may be eukaryotic or prokaryotic. Said transformed host cells include, but are not necessarily limited to, microorganisms, such as bacteria, which have been transformed with a bacteriophage or recombinant plasmid; a yeast transformed with a recombinant vector; animal cells, such as mammalian cells, infected with a recombinant virus vector, eg, adenovirus or vaccinia virus, among others; and insect cells transformed with a recombinant virus vector, e.g., a baculovirus vector. For the expression and harvest of fusion proteins in vitro, bacterial cells can be used as host cells. For example, an E. coli strain, such as, eg, strain DH5 available from the ATCC, Rockville, MD, USA, may be used. (accession number 31343 to the ATCC) or from Stratagene (La Jolla, CA) or strain BL21 available from microorganism deposits such as the ATCC. Eukaryotic host cells, including yeast cells and vertebrate cells, e.g., derived from a mouse, hamster, cow, monkey or human cell line, among others, can also be effectively used. Examples of eukaryotic host cells that can be used to express a fusion protein of the invention include Chinese hamster ovary cells (CHO from Chínese hamster ovary) (e.g., with access CCL-61 number to the ATCC) , Swiss NIH, NIH / 3T3 mouse embryo cells (eg, with accession number CRL-1658 to the ATCC) and Madin-Darby bovine kidney cells (MDBK of Madin-Darby bovine kidney) (CCL number) -22 access to the ATCC). Other cells that are particularly useful for in vitro expression and harvesting of fusion proteins of this invention are cells that possess a system for the glycosylation of amino acids of proteins. Some examples of cells that have a glycosylation system are insect cells, mammalian cells and yeast cells. Systems from different cell types can provide different glycosylation patterns for a fusion protein of the invention. The recombinant vector of the invention is preferably transformed or transfected into one or more host cells of a substantially homogeneous cell culture. The vector can be introduced into host cells according to known techniques, such as, eg, by protoplast transformation, calcium phosphate precipitation, calcium chloride treatment, microinjection, electroporation, transfection by contact with a recombinant virus. , transfection mediated by liposomes, transfection with DEAE-dextran, transduction, conjugation or bombardment with microprojectiles, among others. The selection of transformants can be performed by classical processes, such as selection for cells that express a selectable marker, eg, antibiotic resistance, associated with the recombinant expression vector. Once an expression vector has been introduced into the host cell, integration and maintenance of the polynucleotide sequence encoding a fusion protein of the present invention, either in the genome of the host cell or episomally, can be confirmed by classical techniques, eg, by Southern hybridization analysis, restriction enzyme analysis, PCR analysis [of Polymerase Chain Reaction (including a PCR of reverse transcriptase (rt [reverse transcriptase] -PCR) or by immunological analysis to detect the expected fusion protein product.The host cells which contain a polynucleotide coding sequence and / or which express a fusion protein of The present invention can be identified by any one of at least four general approaches that are well known in the field of technology, including: (i) DNA and DNA hybridization, DNA and RNA or RNA and antisense RNA; (ii) detection of the presence of "marker" gene functions; (iii) determining the level of transcription as measured by the expression of specific mRNA transcripts in the host cell; or (v) detection of the presence of a mature polypeptide product, eg, by immunoassay, as is known in the field of technology. Once a polynucleotide sequence encoding a fusion protein of the present invention has been stably introduced into an appropriate cell, the transformed cell can be clonally propagated, and the resulting cells can be grown under conditions that lead to maximum production of the encoded fusion protein. Such conditions typically include growing transformed cells at a high density. When the expression vector comprises an inducible promoter, suitable induction conditions such as, eg, temperature displacement, nutrient depletion, addition of free inducers (eg, carbohydrate analogs, such as isopropyl- -D-thiogalactopyranoside (IPTG)), accumulation of excess metabolic byproducts, or other similar, are used when necessary to induce an expression.

When the recombinantly expressed fusion protein is retained within the host cells, the cells are harvested and used, and the product is purified from the lysed material under known extraction conditions in the technology sector to minimize protein degradation such as as, for example, at 4 ° C, or in the presence of protease inhibitors, or both at the same time. When the recombinantly expressed fusion protein is secreted from the host cells, the spent nutrient medium can simply be picked up and the fusion protein can be isolated therefrom. The recombinantly expressed fusion protein can be purified from cell lysates or from a culture medium, as necessary, using classical methods, including, but not limited to, one or more of the following methods: Ammonium sulfate, size fractionation, ion exchange chromatography, HPLC, density centrifugation, and affinity chromatography. The recombinantly expressed fusion protein can be detected based, for example, on the size or reactivity with an antibody specific for a fusion protein, or the presence of a fusion tag, eg, a tag. 6XHIS. The present invention encompasses a fusion protein expressed recombinantly in an unpurified state, such as has been secreted into the culture fluid or as is present in a cell lysate material, as well as a partially or substantially purified recombinant fusion protein, all of them being useful for putting the present invention into practice.

Vaccines, including Double Function Vaccines, and Methods for Use The fusion proteins, vectors and transformed cells of the present invention can be used to prepare double-function vaccines for the purpose of inducing an immune suppressive response in a vertebrate against the peptide to which portion (a) of analogue is analogous. the fusion proteins in question, while simultaneously protecting against an infection by the pathogen from which the portion (b) is derived. Such vaccines are also useful in a vertebrate only by inhibiting a peptide to which portion (a) is analogous. Therefore, in one aspect, this invention provides a double-function vaccine comprising a fusion protein as described above, or a vector or a transformed cell comprising a polynucleotide molecule comprising a nucleotide sequence encoding said fusion protein, in an amount effective to inhibit the activity of the peptide from which the portion (a) is derived and to protect with respect to an infection by the pathogen from which the portion (b) is derived in a vertebrate that synthesizes endogenously the peptide and that can be pathogenically infected by the pathogen, together with an acceptable support for pharmaceutical or veterinary use. In a preferred embodiment, the present invention provides a double-function vaccine for inhibiting GnRH activity in cattle while simultaneously protecting this cattle with respect to an infection with BHV-1, which comprises a fusion protein. according to the present invention, or a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, wherein the (a) portion of the fusion protein is analogous all or a part of a GnRH peptide and in which portion (b) is analogous to all or a portion of an antigen of BHV-1, the fusion protein being present in an amount effective to inhibit the activity of GnRH in cattle and also to protect cattle with respect to an infection with BHV-1, together with an acceptable support for veterinary use. The present invention also provides a method for inhibiting the activity of a peptide synthesized endogenously in a vertebrate and for protecting the vertebrate with respect to a pathogenic infection., which comprises immunizing the vertebrate with an amount of a double-function vaccine as described above, which amount is effective to inhibit the activity of the peptide and to protect it from infection by the pathogen. In a preferred embodiment, the present invention provides a method for inhibiting sexual characteristics and for protecting against an infection with BHV-1 in a cow, which comprises vaccinating the cow with a double-function vaccine as before. described, comprising a fusion protein which in turn comprises a portion (a) analogous to all or a portion of a GnRH peptide and a portion (b) analogous to all or a portion of an antigen of BHV- 1, or a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, in an amount effective to inhibit sexual characteristics and to protect against infection by the BHV-1. In vaccines comprising a fusion protein of the invention in which portion (b) is analogous to all or a portion of an antigen of BHV-1, the vertebrate that is vaccinated does not need to be a vertebrate to which the BHV- 1 is able to infect pathogenically. In such vertebrates, portion (b) acts simply as a support to induce an immune response that inhibits the peptide with which it is connected. Therefore, the present invention also provides a vaccine for inhibiting the activity of a peptide in a vertebrate comprising a fusion protein of the invention wherein the portion (a) is analogous to all or a portion of a peptide and the portion (b) is analogous to all or a portion of a BHV-1 antigen, or a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, an amount effective to inhibit the activity of the peptide, together with an acceptable support for pharmaceutical or veterinary use. In a preferred embodiment, the invention provides a vaccine for inhibiting GnRH activity in a vertebrate, comprising a fusion protein in which portion (a) is analogous to all or a portion of a GnRH peptide and the portion ( b) is analogous to all or a portion of a BHV-1 antigen, or a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, in an amount effective to inhibit the activity of GnRH, together with an acceptable support for pharmaceutical or veterinary use. The present invention also provides a method for inhibiting the activity of a peptide, including, but not limited to, the hormone GnRH, in a vertebrate, which comprises immunizing the vertebrate with a vaccine amount described above comprising a fusion, or a vector or a transformed cell comprising a polynucleotide molecule comprising in turn a nucleotide sequence encoding said fusion protein, which fusion protein comprises a protein portion analogous to all or a portion of an antigen of the BHV-1 as a support, whose amount is effective to inhibit the activity of the peptide. The present invention also provides a method for inhibiting sexual characteristics in a vertebrate, preferably a mammal, comprising immunizing the vertebrate with an amount of a vaccine comprising a fusion protein which in turn comprises a portion (a) analogous to all or a part of a GnRH peptide and a portion (b) analogous to all or a portion of an antigen of BHV-1, or a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, whose amount is effective to inhibit sexual characteristics. The vertebrate does not need to be a member of the bovine species, but can be any vertebrate in which the GnRH is endogenously synthesized such as a lamb, a pig, a horse, a goat, a dog, a cat or a human being. "Sexual characteristics" refers to the characteristics present in a vertebrate that are associated with the sexual gender of the vertebrate and / or with the ability of the vertebrate to reproduce, whose characteristics are induced, either in whole or in part, or directly or indirectly, by the GnRH. These characteristics can be determined by those who have ordinary experience in the technology sector. In male animals of cattle, examples of an inhibition of said sexual characteristics include repression of aggressive behavior, suppression of testosterone production, reduced libido, regression of accessory sex glands (including prostates and seminal vesicles), decrease in testicular volume, and reduction or cessation of spermatogenesis. In female cattle animals, the inhibition of said sexual characteristics include failure in ovulation and infertility, regression of the reproductive tract, and abortion. In one embodiment, GnRH is inhibited in a vertebrate either male or female in such a way that the sexual characteristics that are inhibited include a functional reproductive system, the present invention therefore providing a form of contraception. The present invention also provides a method for inhibiting abnormal cell growth in prostate tissue in a male vertebrate, preferably in a mammal, which comprises immunizing the vertebrate with a certain amount of a vaccine comprising a fusion protein, or a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, which fusion protein comprises a portion (a) analogous to all or a portion of a GnRH peptide and a portion (b) analogous to all or a part of an antigen of BHV-1, the amount of which is effective to inhibit the abnormal growth of prostate cells. The vaccines of the present invention may be formulated following accepted conventions to include carriers (carriers) acceptable to animals, including humans, such as classical buffers, stabilizers, diluents, preservatives and / or solubilizers, and may also be formulated to facilitate the release prolonged The diluents include water, a saline solution, dextrose, ethanol, glycerol and the like. Additives to obtain isotonicity include sodium chloride, dextrose, mannitol, sorbitol and lactose, among others. The stabilizers include albumin, among others. Other suitable vehicles and additives for vaccines, including those which are particularly useful for formulating modified live vaccines, are known or will be apparent to those skilled in the art sector. See, eg, the work Remington's Pharmaceutical Science, 18th edition, 1990, Mack Publishing, which is incorporated herein by reference. The vaccines of the present invention may further comprise one or more additional modulatory components of immunity such as, for example, an adjuvant or a cytokine, a cholera toxin (Cholera toxin CT) or a heat labile toxin (LT). de a¿ »/ 7e toxin), among others. Non-limiting examples of adjuvants that can be used in the vaccine of the present invention include the adjuvant system of RIBI (Ribi, Inc., Hamilton, MT), alum, mineral gels such as an aluminum hydroxide gel, emulsions of the type of oil in water, water-in-oil type emulsions such as, eg, Freund's complete and incomplete adjuvants, a block copolymer (CytRx, Atlanta, GA), QS-21 (Cambridge Biotech Inc., Cambridge, MA), SAF-M (Chiron, Emeryville CA), the adjuvant AMPHIGEN®, saponin, Quil A or other saponin fraction, monophosphoryl lipid A, and the lipid adjuvant -Avridine amine. Non-limiting examples of emulsions of the oil in water type, useful in the vaccine of the invention, include the modified formulations SEAM62 and SEAM1 / 2. Modified SEAM62 is an oil-in-water type emulsion containing 5% (v / v) of squalene (Sigma), 1% (v / v) SPAN® 85 (ICI Surfactants) detergent, 0.7% (v / v) of the TWEEN® detergent (ICI Surfactants), 2.5% (v / v) of ethanol, 200 g / ml of Quil A, 100 g / ml of cholesterol and 0.5% (v / v) of lecithin . The modified SEAM 1/2 is an oil-in-water type emulsion comprising 5% (v / v) of squalene, 1% (v / v) of the SPAN® 85 detergent, 0.7% (v / v) of the Tween® 80 detergent, 2.5% (v / v) ethanol, 100 g / ml Quil A and 50 g / ml cholesterol. Other immunomodulatory agents that may be included in the vaccine include, eg, one or more ether ternins, interferons or other known cytokines. When the vaccine comprises living transformed cells, the adjuvant is preferably selected based on the ability of the resulting vaccine formulation to maintain at least some degree of viability of the transformed transformed cells. A vaccine comprising transformed cells of the present invention can be prepared by classical techniques, for example using an aliquot of a culture fluid containing said transformed cells, either free in the medium or resident in mammalian host cells, or both. things at the same time, that can be administered directly, or in a concentrated way, to the individual. Alternatively, modified live transformed cells can be combined with an acceptable carrier for pharmaceutical or veterinary use, with or without an immunomodulatory agent, selected from those that are known in the field of technology and appropriate for the route of administration that has been chosen, wherein at least a certain degree of viability of the living cells is maintained in the vaccine composition. Said methods are known in the art sector. When a vaccine of this invention comprises living transformed cells, the vaccine can be stored in the cold or in the frozen state. When the vaccine composition comprises a fusion protein, a vector, or deactivated transformed cells of the present invention, the vaccine may be stored in the frozen state, or in a lyophilized form to be rehydrated before administration using an appropriate diluent. The vaccines of the present invention may optionally be formulated for the prolonged release of the fusion protein. Examples of such sustained release formulations include the fusion protein in combination with biocompatible polymer composite materials, such as, eg, poly (lactic acid), poly (lactic-co-glycolic acid), methylcellulose, hyaluronic acid, collagen and similar materials. The structure, selection and use of degradable polymers in drug delivery supports have been compiled in several publications, including that of A. Domb et al., 1992, Polymers for Advanced Technologies 3: 279-292, which is incorporated into the present for your reference. An additional guide for selecting and using polymers in pharmaceutical formulations can be found in the text by M. Chasin and R. Langer (editing coordinators), 1990, "Biodegradable Polymers as Drug Delivery Systems" in: Drugs and the Pharmaceutical Sciences, volume 45 , M. Dekker, NY, which are also incorporated herein by reference. Alternatively, or additionally, the fusion protein, vector or transformed cells can be microencapsulated to provide administration and efficacy. Methods for microencapsulating antigens with well-known in the technology sector, and include the techniques described, e.g., in U.S. Pat. 3,137,631; in U.S. Pat. 3,959,457; in U.S. Pat. 4,205,060; in U.S. Pat. 4,606,940; in U.S. Pat. 4,744,933; in U.S. Pat. 5,132,117; and in International Patent Publication WO 95/28227, all of which are incorporated herein by reference. Liposomes can also be used to provide prolonged release of the fusion protein, the vector or the transforcell. Details concerning how to produce and use liposome formulations can be found, among other citations, in U.S. Pat. 4,016,100; in U.S. Pat. 4,452,747; in U.S. Pat. 4,921,706; in U.S. Pat. 4,927,637; in U.S. Pat. 4,944,948; in U.S. Pat. 5,008,050; and in U.S. Pat. 5,009,956, all of which are incorporated herein by reference. An effective amount of any of the vaccines described above can be determined by conventional means, starting with a low dose of a fusion protein, a vector or a transforcell, and then increasing the dosage while monitoring the effects. An effective amount can be obtained after a single administration of a vaccine or after multiple administrations of a vaccine. Known factors can be taken into consideration when determining an optimal dose per animal. These include the species, size, age and general condition of the animal, the presence of other drugs in the animal, and similar factors. The actual dosage is preferably chosen after taking the results from other animal studies into consideration. One method of detecting if an adequate immune response has been achieved is to determine the seroconversion and the antibody titer in the animal after a vaccination. The timing of the vaccination and the number of revaccinations, if any, will preferably be determined by a qualified scientist or veterinarian based on the analysis of all relevant factors, some of which have been described above. The amount of effective dose of a fusion protein, a vector and a transforcell of the present invention can be determined using known techniques, taking into account factors that can be determined by a person having ordinary experience in the technology sector, such as the weight of the animal to be vaccinated. The dose amount of the fusion protein of the present invention in a vaccine of the present invention ranges preferably between about 1 g and about 10 mg, more preferably between about 50 g and about 1 mg, and most preferably between about of 100 g and about 0.5 mg. The dose amount of a vector of the present invention in a vaccine of the present invention preferably ranges from about 50 g to about 1 mg. The number of doses of transforcells of the present invention in a vaccine of the present invention preferably ranges from about 1 x 10"3 to about 1 x 10" cells / ml, and more preferably from about 1 x 10 ^ and about 1 x 10 'cells / ml. An appropriate dosage amount ranges from about 0.5 ml to about 10 ml and more preferably between about 1 ml and about 5 ml. When the inhibition of abnormal cell growth in the prostate is involved, an effective amount of any of the above-described vaccines can be determined by conventional means, starting with a low dose of a fusion protein, a vector or a transformed cell, and then increasing the dosage while the effects are being monitored.

Known factors can be taken into consideration when determining an optimal dose per animal. Some of the factors have been described above. "Abnormal cell growth" means cell growth that is independent of normal regulatory mechanisms (e.g., loss of contact inhibition). This includes the abnormal growth of (1) cells from malignant prostate tumors, such as prostate carcinoma cells, (2) benign cells from other proliferation disorders in a prostate tissue, and (3) any other unregulated growth. of cells in a prostate tissue associated with GnRH activity. "Inhibiting the growth of prostate carcinoma" and similar phrases as used in the present context mean decelerating, stopping and / or reversing the abnormal growth of cells in a prostate tissue. The present invention further provides a method of preparing a vaccine comprising a fusion protein as described above, which method comprises combining an effective amount of a fusion protein of the present invention with a support acceptable for pharmaceutical or veterinary use.

Antibodies The present invention further provides a method for producing monoclonal antibodies, directed against a peptide that is endogenously synthesized in a vertebrate, comprising vaccinating said vertebrate with an antibody-inducing amount of a fusion protein of the present invention, or a vector or a transformed cell comprising a polynucleotide molecule comprising in turn a nucleotide sequence encoding said fusion protein, which fusion protein comprises a portion (a) analogous to all or a portion of a peptide endogenously synthesized within the vertebrate; obtain a serum containing polyclonal antibodies from the vaccinated vertebrate; and isolating from the serum the polyclonal antibodies that bind to the peptide that has been synthesized endogenously; thus producing polyclonal antibodies directed against the peptide. The methods for obtaining a serum from a vaccinated vertebrate and for isolating specific polyclonal antibodies from it are known in the field of technology. In a preferred embodiment, the fusion protein comprises a portion (a) analogous to all or a part of a GnRH peptide, and the peptide against which polyclonal antibodies are produced is GnRH. The present invention further provides polyclonal antibodies directed against an endogenously synthesized peptide that has been produced according to this method. In a preferred embodiment, the polyclonal antibodies are directed against GnRH. The present invention further provides a method for producing a monoclonal antibody directed against a peptide that is endogenously synthesized in a vertebrate, comprising vaccinating said vertebrate with an antibody-inducing amount of a fusion protein of the present invention, or a vector or a transformed cell comprising a polynucleotide molecule which in turn comprises a nucleotide sequence encoding said fusion protein, which fusion protein comprises a portion (a) analogous to all or a portion of a peptide synthesized endogenously within the vertebrate; and isolating a spleen cell from the vaccinated vertebrate whose spleen cell excretes a monoclonal antibody that binds specifically to the endogenously synthesized peptide; thus producing a monoclonal antibody directed against the peptide. In a preferred embodiment, the fusion protein comprises a portion (a) analogous to all or a part of a GnRH peptide, and the peptide against which the monoclonal antibody is produced is GnRH. The present invention further provides monoclonal antibodies directed against an endogenously synthesized peptide that has been produced according to this method. In a preferred embodiment, the monoclonal antibodies are directed against GnRH. Methods for isolating spleen cells from a vaccinated animal that excretes a specific monoclonal antibody for the purposes of producing a monoclonal antibody are known in the field of technology. Such methods include, but are not limited to, the hybridoma technique originally described by Kohier and Milstein (Nature, 1975, 256: 495-497); the human B-cell hybridoma technique (Kosbor et al., 1983, Immunology Today 4:72; Cote et al., 1983, Proc. Nati Acad. Sci. USA 80: 2.026-2.030); and the EBV hybridoma technique [Epstein-Barr virus] (Colé et al., 1985, Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pages 77-96). These publications are incorporated herein by reference. Techniques for the production of monoclonal antibody fragments and antibodies are further described, inter alia, in Harlow and Lane, 1988, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, and in that of JW Goding, 1986, Monoclonal Antibodies: Principies and Practice, Academic Press, London, which are incorporated herein by reference. The following Examples are provided to illustrate merely aspects of the present invention. These are not intended, and should not be considered, as limiting the invention set forth in the claims and described more fully in this specification.

EXAMPLES EXAMPLE 1 Plasmids expressing fusion proteins of gD and GnRH Construction of pQE-tmgD: Plasmid FlgD / Pots207nco (# 79) (which encodes a full length gD, hereinafter referred to as "gD / Pots") was digested with Ncol and Xbal and the resulting fragment of 1.26 kb it was cloned into the corresponding pUC21 sites, generating the plasmid pUC-FLgD. The complete sequence of the Ncol and Xbal fragment in the plasmid pUC-FLgD was determined on both strands of DNA using Sanger's fluorescent dideoxy chain termination sequencing technology. Figure 3 shows the results and the characteristics of the sequences. The nucleotide sequence encoding gD / Pots is included in SEQ ID NO: 16. The alignment of the DNA between the gD / Pots and the gD of the published BHV-1 (accession number M59846 to GenBank) has a homology of 94, 7% with most of the mismatches that occur in 3 'of the transmembrane domain (Figure 4). The amino acid alignment between gD / Pots and M59846 shows four differences between amino acids, one of which is located in the signal sequence and the other three are located in or around the transmembrane domain (Figure 5). The signal sequence for the gD protein was deleted in order to facilitate the expression of the protein in E. coli. The deletion of the signal sequence was carried out by digesting the plasmid DNA pUC-FLgD with Ncol and HindIII and filling the ends with the Klenow fragment of DNA Polymerase I. The resulting DNA fragment was gel purified and ligated. The colonies were screened for a shift in mobility of the Sphl and Sacl fragment that would indicate deletion of the 50 bp fragment (base pairs). Two positive clones were selected and sequenced through the deletion region with Ncol and Hindlll. It was shown that all the clones have the correct sequence. Clone # 1 was designated pUC-MgD and was chosen for further manipulation. The mature sequence of gD was subcloned into an expression vector in E. coli, for production of the protein on a large scale basis. To this end, a 1. 07 kb Sphl and Sacl fragment from pUC-MgD containing the mature gD sequence (truncated at the 3 'end to exclude the transmembrane domain) was subcloned into the corresponding pQE- sites. 31 (Qiagen). (PQE-31 uses the phage T5 promoter and two lac operator sequences for further repression prior to induction of expression with IPTG.) PQE-31 also contains a 6XHIS-tag fusion at the terminal end of N for purification purposes). The resulting clones were screened for the Sphl and Sacl fragment of 1. 07 kb. A positive clone, designated pQE-tmgD, was selected for the preparation of additional plasmids, see below. The pQE-tmgD encodes a 6XHIS tag at the N-terminus fused to a truncated mature gD sequence (tmgD), terminated by a stop codon encoded by a vector following the site for Sacl. The binding regions of the gD sequence and the plasmid framework were sequenced to verify the integrity of the insert, and found to be correct. The sequence encoding the tmgD (which does not include this 6XHIS tag) in the pQE-tmgD is set forth in SEQ ID NO: 36. The amino acid sequence of the tmgD encoded by the pQE-tmgD (without the 6XHIS tag) is set forth in SEQ ID NO: 35 Construction of the GnRH tetramer clones Twelve different oligonucleotides (reverse sense and complementary strands) that encoded GnRH (monomers and dimer) having different terminal DNA sequences were prepared. These twelve oligonucleotides are presented in SEQ ID NOS: 1-12. Oligonucleotides 9 and 10 were annealed and cloned into the BamHI and XhoI sites of pBS KS + (Stratagene), generating p98BS / GnRH. A plasmid encoding a GnRH tetramer was constructed from plasmid p98BS / GnRH by adding oligonucleotides 7 and 8 annealed at the sites for SmaI and XhoI. A full length tetramer reconstruction was necessary, since analysis of the sequences of 5 separate clones showed that all of them had base changes in the region of synthetic primer. A clone containing the full length tetramer with the correct sequence was constructed by replacing a 106 bp Eagl fragment from a mutant clone with the corresponding fragment from a clone lacking base changes in this region. One of the resulting reconstructed clones was sequenced and found to have the correct DNA sequence encoding the GnRH tetramer. This clone contained a difference between sequences with respect to the predicted nucleotide sequence for the artificial structure of the GnRH tetramer. The change is an additional G at 3 'and outside the coding region of GnRH and, therefore, does not affect the coding region for GnRH. (The additional G was present in the clone used for the reconstruction, and it is probably due to an error in the sequence of the synthetic primer). This clone was designated as p9897-R. A portion of p9897-R, including the sequence encoding the GnRH tetramer, is shown in Figure 2. The sequence encoding the GnRH tetramer is set forth in SEQ ID NO: 32. The amino acid sequence of the encoded GnRH tetramer for p9897-R is set forth in SEQ ID NO: 33. A PCR was used using the primers P14-S1 (SEQ ID NO: 42) and P14-A138 (SEQ ID NO: 43) with a template DNA from the plasmid p9897 -R to generate a 138 bp fragment containing the PCR fragment of the GnRH tetramer having a 3 'stop codon and ends for Sacl in 5' and for HindIII in synthetic 3 '. The PCR fragment was cloned into the eASY pGEM-T vector (Promega, Madison, Wisconsin), generating p9897 S / d3. The clone was sequenced and found to have the correct sequence. The clone, p9897 S / d3, provides a source for a coding sequence of the GnRH tetramer with ends for Sacl and Hindlll for future cloning into pQE vectors.

Construction of pQE-cD: GnRH A 126 bp Sacl / Hindlll fragment from p9897 S / d3 containing the GnRH tetramer was cloned into the corresponding sites of the pQE-tmgD plasmid. The colonies were screened for the presence of the Sacl and Hindlll fragment of 126 bp and a fragment of BamHI and Hindlll of 1165 bp indicating an appropriate orientation of the insert. The binding regions adjacent to the cloning sites were analyzed by DNA sequencing and found to be correct. The nucleotide sequence encoding tmgD-4GnRH, which includes the 6XHIS tag and flanking sequences of the plasmid is set forth in SEQ ID NO: 24. The amino acid sequence of the tmgD-4GnRH, encoded by pQE-gD: GnRH is set forth in SEQ ID NO: 25. As previously described, tmgD-4GnRH is an artificial fusion structure in which a GnRH tetramer is fused to the terminal carboxy terminus of the truncated mature gD.

Construction of pQE-GnRH: gD The coding sequence of the GnRH tetramer in p9897-R was cut with BamHI and Ncol, the ends blunted by filling in with Klenow and the 132 bp fragment was gel purified. A mature gD vector fragment (ie without the signal sequence) was prepared by cutting from pUC-FLgD with Ncol and Hindlll, blunting the ends by filling them with Klenow, and gel-purifying the 4.4 kb fragment . After ligation with the 132 bp fragment from p9897-R and transformation, the clones were screened for regeneration of the 5 'BamHI and Ncol sites resulting from ligation in the correct orientation. An additional scrutiny regarding the generation of a fragment of Ndel of ~ 400 bp, confirmed the correct structure. The artificial structure was sequenced through the junctions between GnRH and gD to confirm the correct sequence. This artificial structure, designated pUC-GnRH: gD, contains a GnRH tetramer sequence fused to the amino terminal end of a full-length mature gD sequence in a pUC vector. A fusion sequence of the GnRH tetramer and the truncated mature gD was obtained by digesting the pUC-GnRH: gD with the restriction enzymes Sphl and Sacl. This 1161 bp fragment was cloned into the corresponding sites of pQE-31, generating the pQE-GnRH: gD. The clones were screened for the Sphl / Sacl fragment of 1161 bp and for the correct model of the Ndel fragments (380 bp, 2.0, 2.2 kb). The nucleotide sequence encoding the 4GnRH-tmgD, including the 6XHIS tag, and the plasmid flanking sequences are set forth in SEQ ID NO: 22. The amino acid sequence of the 4GnRH-tmgD encoded by the pQE-GnRH: gD is exposed in SEQ ID NO: 23.

Construction of pQE-GnRH: aD: GnRH The 126 bp Sacl / Hindlll fragment from p9897 S / d3 was subcloned into the corresponding sites of plasmid pQE-GnRH: gD, generating pQE-GnRH: gD: GnRH. The clones were screened for the 126 bp Sacl / Hindlll fragment as well as for the correct fragment model for Ndel. PQE-GnRH: gD: GnRH encodes a fusion protein 4GnRH-tmgD-4GnRH. As previously described, 4GnRH-tmgD-4GnRH comprises a truncated mature gD having a GnRH tetramer fused at both amino and carboxy ends. The nucleotide coding sequence and the flanking sequences from pQE-GnRH: gD: GnRH are presented in SEQ ID NO: 26. The amino acid sequence of 4GnRH-tmgD-4GnRH, encoded by pQE-GnRH: gD: GnRH, including the 6XHIS brand, is set forth in SEQ ID NO: 27.

Comparison of expression products from artificial structures with the pQE vector of bacterial expression. All four artificial structures contained a tmgD derived from clone FlgD / Pots207nco (# 79), which included amino acids 19 to 358 of FlgD / Pots207nco (# 79). All four artificial structures contained a leader sequence of pQE-HIS at the terminal end of amino (a 6XHIS tag) designated by the amino acid designation: MRGSHHHHHHTDPHA (SEQ ID NO: 37). The coding sequence for the 6XHIS tag is set forth in SEQ ID NO: 38. All four artificial structures had a 2 or 3 amino acid linker behind the 6XHIS leader sequence. All GnRH products were derived from a p9897-R clone of the GnRH tetramer. The clones pQE-GnRH: gD and pQE-GnRH: gD: GnRH contained a three amino acid linker (SMS) between the GnRH tetramer at the amino terminal end and the tmgD sequence. The clones pQE-gD and pQE-GnRH: gD contained about ten additional amino acids at the terminal carboxy terminus of tmgD from the vector sequence as an aberration due to cloning. The clones pQE-gD: GnRH and pQe-GnRH: gD: GnRH contained a linker with an amino acid (proline) between tmgD and the fusion of carboxy and GnRH. See Figure 10 for an illustration of each of the artificial pQE structures.

EXAMPLE 2 Expression of GnRH and gD Fusion Proteins by Transformed Bacterial Cells All of the artificial pQE structures described in Example 1, above, were transformed into the cells of DH5-F'IQ cells of E. coli for their expression. For the induction of expression, the cells were grown to an OD600 of 0.7-0.9 in a culture flask fitted with 2 liter deviating elements in a 2xYT broth containing 100 g / ml of ampicillin and 25 g / ml kanamycin, then induced with 1-2 mM IPTG and incubated for 4 hours at 37 degrees Celsius. DO readings 500 average at the time of harvest were 1, 3. The expression of all four artificial structures was confirmed by Western blot analysis.

EXAMPLE 3 Formulation of Vaccines with Fusion Proteins and Immunization of Mice Assemble of vaccines The fusion proteins from pQE-tmgD (as a control), pQE-GnRH: gD, pQE GnRH: gD: GnRH and pQE-gD: GnRH were concentrated from preparations of inclusion bodies by preparative electrophoresis on 9% polyacrylamide gels. Bands cut from the SDS PAGE gels were dissolved in 25 mM Tris, pH 8.3, 192 mM glycine and 0.1% SDS (w / v). The equivalent of a 10 g dose of gD / mouse was adjuvanted with a SEAM1 emulsion (Squalene Emulsion Adjuvant Metabolizable = metabolizable by emulsion adjuvant with squalene) (10 g dose of Quil A / 100 I. The vaccine formulations were stored at 4 C. SEAM1 is 5% squalene, 0.1% vitamin E acetate, 1% Span 85, 0.70% Tween 80, 2 mg / ml QuilA and 400 l / ml cholesterol.

Male BALB / c mice were used in the study after they were 8 weeks old (10 per group). The mice were initially housed in groups of 10, but the controls were subsequently moved to individual cages to prevent them from fighting.

Immunization Mice were immunized subcutaneously with 10 g of a fusion protein in 100 I of an adjuvant, as described above. Three immunizations were administered on days 0, 20 and 41 of the study.

Anti-GnRH antibodies by ELISA Samples were collected from sera on days 0, 20, 31, 41, 55, 62, 69 and 146 of the study and were evaluated for anti-GnRH antibody titers in an ELISA (analysis by enzyme linked immunosorbent = enzyme linked immunoadsorbant assay) of peptides. A biotinylated GnRH peptide (Biotin-GnRH) (0.1 g / ml in 25 mM Tris, 0.15 M NaCl at pH 7.6) consisting of the natural sequence plus a 4 amino acid linker (CAGAEHWSYGLRPG), purified by HPLC on a reverse phase column, adsorbed to avidin-coated plates and incubated at room temperature for 2 hours. The excess peptide was removed by washing the plates four times with the wash buffer (25 mM Tris, 0.15 M NaCl, 0.05% Tween-20 and BSA (bovine serum albumin = fraction of bovine serum) fraction V) . Then, serial, quintuple dilutions of sera from positive, negative and unknown control mice were added in a diluent (25 mM Tris, 0.15 M NaCl, 0.05% BSA) (100 μl / well) to the wells coated with the peptide and incubated for 30 minutes at room temperature. The plates were washed four times in a wash buffer and then a rabbit anti-IgG mouse horseradish peroxidase (IgG-specific) horseradish peroxidase (Zymed, California) (1: 4000, 100 l / well) was added to each well. . After incubation for 30 minutes at room temperature, the bound antibody was detected with a substrate of 3,3 ', 5,5'-tetramethyl-benzidine (Kierkegaard &Perry, Catalog No. 50-76-04) ( 100 μl / well, for 15 minutes in the dark) and the reaction was stopped by the addition of 50 μl / well of 0.18 M H2SO4. The absorbance at 450 nm was measured with a microplate reader from Molecular Devices. To calculate the antibody titers, a positive control curve is generated and the titres of the unknown samples are extrapolated from the curve using a computer logic system.

ELISA for BHV-1 oD Serum samples were collected on days 0, 20, 31, 41, 55, 62, 69 and 146 of the study and were evaluated for antibody titers of anti-gD BHV-1 by an ELISA The recombinant gD of BHV-1, purified and expressed from MDBK cells (Madin Darby Bovine Kidney - from Madin Darby bovine kidney (1 g / ml in Dulbecco PBS + 0.01% thimerosal, 100 I / well) was adsorbed on microtiter plates for 18-24 hours at 4 C. The excess protein was washed from the wells and then the unfixed sites in the wells were blocked by incubating for 2 hours at 37 C with 300 I of PVA (polyvinyl acetate) in 1% DPBS (Dulbecco's phosphate buffered saline = Dulbecco's phosphate buffered saline solution) with 0.01% thimerosal. Serum samples (positive and negative and unknown control sera) were diluted 1: 50, then serially by quadruple dilutions in 1% PVA in DPBS with 0.01% thimerosal and 100 μl was added to each well. The test material was incubated for 45 minutes at 37 C. The plates were washed four times with distilled H2O, then HRP (horseradish peroxidase = horseradish peroxidase) of goat anti-mouse (1: 10,000 in PVA 1 was added). % in DPPBS with 0.01% thimerosal, 100 l / well, KP + L) and the plates were incubated for 30 minutes at 37 C. The wells were washed four times with distilled H20 and then the analysis material was developed with the substrate of ABTS (2,2, -azino-di [3-ethyl-benzothiazoline-sulfonate (6)] (100 μl / well, RT (room temperature, 15 min.) The reaction material was read at 405/490 nm in an ELISA reader Titers were calculated using the Forecast method in EXCEL® (Microsoft, Redmond, Washington) using an OD of 0.5 as a cut and using 2 dilutions above the OD 0.5 and a dilution per under OD 0.5 to extrapolate the titles.

Testosterone concentrations Samples of sera from days 0, 41 and 69 of the study were evaluated for testosterone concentrations. The analysis was a radioimmunoassay of human testosterone using an antibody that cross-reacts with murine testosterone. Human testosterone patterns are used in the analysis. The murine samples tend to move at the lower end of the human testosterone standard curve, leading to a wider variability in normal values. The sensitivity of the analysis is 0.02 ng / ml.

Necropsy and Histopathology The animals were sacrificed on day 146 of the study. The testicles, the epididymis and the prostate were removed together with the seminal vesicle, and weighed before the fixation of tissues in Bouin's fixative agent [75 ml of picric acid (saturated solution); 25 ml of formalin (37%), 5 ml of acetic acid (4.76%)]. The tissues were fixed for 48 hours and then rinsed in a 50% mixture of ethanol and H2O. The tissues were stored in fresh 50% ethanol before analysis. Tissues were treated and embedded in paraffin and sections of 5 m were cut and stained with hematoxylin and eosin. Each of the organs was evaluated for inflammation, atrophy and spermatogonial degeneration. Grades were assigned based on the level of spermatogenesis, atrophy or other lesions. The weights were scored as a percentage of the average weight in the normal control group. An accumulated score was assigned to each animal.

Results Anti-gD antibody responses All mice that had been immunized with gD or with a fusion protein containing gD generated anti-gD ELISA antibodies, regardless of whether the gD was expressed in prokaryotic expression systems (i.e. carboxyl, amino or carboxyl-amino fusion expressed in E. coli) or in eukaryotic expression systems (i.e., a protein expressed in MDBK).

Responses of anti-GnRH antibodies Hierarchies of anti-GnRH titers were induced by the different fusion proteins: the tmgD-4GnRH (meaning that it had a GnRH tetramer at the carboxy end of the protein) generated the highest titers, followed by those of the 4GnRH-tmgD-4GnRH, while the lowest titers were induced in the immunized 4GnRH-tmgD. In all groups, the anti-GnRH titers showed a peak (maximum) after the second immunization and remained at a stable value (plateau) for more than 2 months. All mice (9 of 9) immunized with tmgD-4GnRH produced antibody responses to GnRH when measured by a peptide ELISA, although 2 of 9 mice were low responders. There were 3 of 10 non-responders in the 4GnRH-tmgD group and 1 of 9 non-responders in the 4GnRH-tmgD-4GnRH group. All non-responders to GnRH were responders to gD.

Effect of anti-GnRH antibodies on the male reproductive system To determine whether the induction of anti-GnRH antibodies would negate the function of GnRH, we have evaluated testosterone levels before and after immunization with GnRH. When performing the necropsy, the tissues of the reproductive tract were weighed and then subjected to gross and histological examination. The normal margins of testosterone concentrations in mice varied widely when measured using human testosterone radioimmunoassay. However, mice immunized with tmgD-4GnRH had significantly lower mean testosterone concentrations when compared with normal controls or with other treatment groups. The prostate, testes and epididymides of mice immunized with tmgD-4GnRH were significantly atrophied when the raw tissue weight and histological examination of sperm development were evaluated. Mice that had been immunized with 4GnRH-tmgD-4GnRH were less affected when compared to normal controls.

EXAMPLE 4 Artificial baculovirus structures encoding gD v GnRH fusion proteins Construction of pBacHIS D.LH and bac-qD: GnRH The pQE-gD: GnRH (see Example 1) was digested with HindIII, the site blunted by treatment with Klenow, and subsequently digested with EcoRI. A fragment of approximately 1.2 kb containing the coding sequence of tmgD-4GnRH was gel purified and cloned into the transfer vector plasmid pBacPAKT digested with STUI / EcoRI (Clontech Inc.), forming the pBacHISgD ± H. (The transfer vector contains sequences that compensate for the replication deficiency in a baculovirus deficient in replication). Sf21 insect cells were transfected together with pBacHISgD: LH and replication-deficient baculovirus viral DNA. These transfected Sf21 cells generate a recombinant baculovirus (designated as bac-gD: GnRH) (accession number VR-2633 to the ATCC), which encodes a tmgD-4GnRH fusion protein. The recombination (DNA exchange) between the transfer vector pBacHISgD: LH and the replication-deficient baculovirus viral DNA is mediated by homologous flanking viral sequences present in pBacPAK9 which allows efficient transfer of the entire expression cassette (sequence encoding tmgD-4GnRH) from pBacHISgD: LH within viral DNA together with the gene or genes that complement the replication deficiency. The recombinant virus can be purified by plaque analysis from infected Sf21 cells. Repeated cycles of Sf21 cell infection and purification can be performed by plaque analysis to obtain a higher concentration of the recombinant virus expressing the fusion protein for large scale production of the fusion protein. The expression of the recombinant artificial structures was confirmed by Western blotting. Infected Sf21 cells can be harvested by centrifugation and transferred at -80 Celsius until treated by recombinant baculoviruses. The nucleotide sequence encoding the ORF (Open Reading Frame = open reading frame for the 6XHIS tag, truncated mature gD and the GnRH tetramer in bac-gD: GnRH is set forth in SEQ ID NO: 39. Nucleotides Nos. 1-45 encode a 6XHIS tag, nucleotides Nos 46-1,074 encode a truncated mature gD of BHV-1, nucleotides Nos. 1,075-1,194 encode a tetramer of GnRH, and nucleotides Nos. 1,195-1,197 are a stop codon. amino acid sequence of the fusion protein encoded by bac-gD: GnRH is the same as the sequence set forth in SEQ ID NO: 25.

Construction of pBacHISMgD An artificial structure of recombinant baculovirus containing gD was generated as a control. Plasmid pCMV-MgD (see Example 5, infra) was digested with Pací and Apal allowing the isolation of a 950 bp fragment containing the majority of the gD gene minus the 5 'end. Plasmid pBacHISgD: LH underwent digestion with Pací and Apal allowing the isolation of a fragment of 5., 6 kb containing the plasmid framework and the 5 'portion of gD. Ligation of the 5.6 kb fragment with the 950 bp fragment generated plasmid pBacHIsMgD containing mature gD truncated in the transfer vector pBacPAC9. Sf21 cells were transfected together with pBacHISMgD and replication deficient virus. These transformed Sf21 cells generate recombinant baculoviruses (designated as Bac-MgD) that encode tmgD. The recombinant virus was purified and stored as described above.

Expression Recombinant baculovirus can be obtained from lysed materials of infected Sf21 cells. The lysed material also contains the fusion protein expressed by the recombinant virus, and the fusion protein can be purified from the lysed material. For example, after lysis with detergents of the cell pellet, the pellet of lysed material from the aforementioned example was solubilized in 8 M urea, 50 mM Tris, pH 7.5 and loaded onto a Ni NTA column; tmgD-4GnRH was eluted in a step gradient of pH. The lysate containing both the recombinant baculovirus and the fusion protein can be stored, for example at -80 C Celsius.

EXAMPLE 5 Plasmid suitable for in vivo expression of gD v GnRH fusion proteins The ß-Gal gene from the vector pCMVβ (Clontech, Inc.) was removed by restriction digestion with EcoRV and NotI and the resulting vector fragment for NotI was isolated by gel electrophoresis. A synthetic linker containing multiple cloning sites (MC) with ends for Notl was cloned into this Notl vector fragment creating pCMV-MC. A truncated gD gene that included the signal sequence was amplified by PCR from FlgD / Pots207nco (# 79) using primers that introduced a site for EcoRV at the 5 'end, a second codon repaired to encode Gln instead of Glu , a stop codon was added behind Pro 337 of the coding sequence, and a site for Kpnl was added at the 3 'end. This 1083 bp PCR fragment was cloned into the pGEM-T EASY vector digested with EcoRV and Kpnl (Promega, Madison, Wisconsin), generating pGEM-T-EASY / gD, and subsequently sequenced by chemical termination conditions with digestion. -deoxi fluorescent in both directions to ensure the integrity of the PCR product. The truncated gD fragment was isolated from clone pGEM-T-EASY / gD by digestion with EcoRV and Kpnl and subcloned into pCMV-MC. The resulting clone, designated pCMV-gD, was verified by restriction enzyme analysis. To construct the pCMV-gD: GnRH (accession number 203370 to the ATCC), the pQE-gD: GnRH was cut with Hindlll, blunt-ended with a Klenow and then digested with Apal. The resulting blunt end and Apal fragment of 1.05 kb containing the tmgD and the GnRH tetramer was isolated. The clone, pCMV-gD, was cut with Smal, followed by Apal, eliminating the region encoding truncated gD. The remaining 3.7 kb pCMV vector fragment containing the signal sequence for gD was isolated and used in a ligation reaction with the 1. 05 kb fragment containing tmgD and the GnRH tetramer. The resulting clone was designated pCMV-gD: GnRH. The ORF coding for tgD-4GnRH, including the signal sequence, from pCMV-gD: GnRH, is set forth in SEQ ID NO: 28. The amino acid sequence of tgD-4GnRH, including the signal sequence, encoded by pCMV-gD: GnRH, is set forth in SEQ ID NO: 29. All patents, patent applications and publications mentioned above are incorporated herein by reference in their entirety.

The present invention is not to be limited in scope by the specific embodiments, which are intended to be simple illustrations of individual aspects of the invention and which functionally equivalent methods and components are within the scope of the invention. Of course, various modifications of the invention, in addition to those shown and described in the present text, will be apparent to those skilled in the art from the foregoing descriptions of the accompanying drawings. It is intended that such modifications fall within the scope of the appended claims.

Deposit of biological materials The following biological material was deposited in the American Type Culture Collection (ATCC) domiciled at 10801 University BIvd., Manassas, Virginia, 20110-2099, USA. on October 22, 1998, and the following access numbers were assigned: Plasmid Accession Number Plasmid pQE-gD: GnRH 98953 Plasmid pCMV-gD: GnRH 203370 Plasmid pQE-GnRH: gD 98954 Plasmid pQE-GnRH: gD: Gr IRH 98955 Vector Accession number baculovirus bac-gD: GnRH VR-2633 All patents, patent applications and publications mentioned above are incorporated herein by reference in their entirety. The present invention is not to be limited in scope by the specific embodiments, which are intended to be simple illustrations of individual aspects of the invention and which functionally equivalent methods and components are within the scope of the invention. Of course, various modifications of the invention, in addition to those shown and described in the present text, will be apparent to those skilled in the art from the foregoing descriptions of the accompanying drawings. It is intended that such modifications fall within the scope of the appended claims.

LIST OF SEQUENCES < 110 > Pfizer Products Inc < 120 > FUSION PROTIINS THAT INCLUDE SUPPORTS THAT CAN INDUCE A DOUBLE IMMUNE RESPONSE < 130 > PC10202A < 140 > < 141 > < 150 > N / A < 151 > 02-17-1999 < 160 > 46 < 170 > PatentlnVer.2.1. < 210 > 1 < 211 > 33 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 1 catggaacac tggtcrtatg gtctgcgtcc ggg 33 < 210 > 2 < 211 > 33 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 2 catggaacac tggtcttatg gtctgcgtcc ggg 33 < 210 > 3 < 211 > 36 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 40C > 3 qa z zqqaac ac ggi-Ctta "ggtctgcgt ccgggc 36 < 210 > 4 < 211 > 36 < 212 > DNA < 213 > Artificial Sequence < 40C > 4 gatcgcccgg acgcagacca -aagaccagt gt.cca 36 < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 210 > 5 < 211 > 76 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 5 gatccatgga gcactggtca tatggtctgc gtccgggtga acattggagc tacggtctac 60 gccccgggtc catggc 76 < 210 > 6 < 211 > 76 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES 4XJ > o tcgagccatg gacccggggc gtagaccgta gctccaatgt tcacccggac gcagaccata 60 egaccagtgc tccatg 76 < 210 > 7 < 211 > 71 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 7 ggggaacact ggtcttatgg cttacggccg ggagagcatt ggagttacgg cctccgtcca 60 ggttccatgg c 71 < 210 > 8 < 211 > 75 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 8 tcgagccatg gaacctggac ggaggccgta actccaatgc tctcccggcc gtaagccata 60 agaccagtgt tcccc 75 < 210 > 9 < 211 > 71 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 9 gatccagagc actggtcata tggtctgcgt ccgggtgaac attggagcta cggtctacgc 60 cccggsgatc c 71 < 210 > 10 < 211 > 71 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 10 cgaggatcc ccggggcgta gaccgtagct ccaatgttca cccggacgca gaccatatga 60 ccagtgctct g 71 < 21O > 11 < 211 > 68 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES < 400 > 11 ggggaacact ggtcttatgg cttacggccg ggagagcatt ggagttacgg cctccgtcca 60 sgggatcc 68 < 210 > 12 < 211 > 72 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: SYNTHETIC OLIGONUCLEOTIDE THAT COMPRISES A SEQUENCE THAT CODIFIES GNRH AND CLONING EXTREMES ccagtgctcc cc 72 < 400 > 12 tcgaggatcc cctggacgga ggccgtaact ccaatgctct cccggccgta agccataaga 60 < 210 > 13 < 211 > 10 < 212 > PRT < 213 > SEQUENCE OF AMINO ACIDS OF GNRH < 400 > 13 Glu His Trp Ser Tyr Gly Leu Arg Pro Gly 1 5 10 < 210 > 14 < 211 > 328 < 212 > DNA < 213 > Artificial Sequence < 220 > < 223 > Description of the Artificial Sequence: Part of the plasmid p9897-R < 400 > 14 acgccasggt tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg cgcgtaatac 60 gac ac-ac agggcgaatt ggagctccac cgcggtggcg gccgctctag aactagtgga 120 tccagagcac tggtcatatg gtctgcgtcc gggtgaacat tggagctacg gtctacgccc 180 cggggaacac tggtcttatg gcttacggcc gggagagcat ggagttacg gcctccgtcc 240 aggttccatg ggctcgaggg ggggcccggt acccagcttt tgttcccttt agtgagggtt 300 aattgcgcgc ttgscgtaat atggtcat 328 < 210 > 15 < 211 > 40 < 213 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: GnRH Tetramer < 4 CC > 15 Glu His Trp Ser Tyr Gly Leu Arg Pro Gly Glu His Trp Ser Tyr Gly 1 5 10 15 Leu Arg Pro Gly Glu His Trp Ser Tyr Gly Leu Arg Pro Gly Glu His 20 - 25 30 Trp Ser Tyr Gly Leu Arg Pro Gly - 35 40 < 210 > 16 < 211 > 1259 < 212 > DNA < 213 > Bovine Herpesvirus 1 < 220 > < 221 > gene < 222 > (1) .. (1259) < 223 > sequence encoding the gD of BHV-1 from the clone FigD / Pots207 (# 79) < 400 > 16 ccatggaggg gccgacattg gccgtgctgg gcgcgctgct cgccgttgcg gtaagcttgc 60 ctacacccgc gccgcgggtg to ggtatacg tcgacccgcc ggcgtacccg atgccgcgat 120 ácaactacac gaacgctgg acactaccg ggcccatacc gtcgcccttc gcagacggcc 180 gcgagcagcc cgtcgaggtg cgctacgcga cgagcgcggc ggcgtgcgac atgctggcgc 240 tgatcgcaga cccgcaggtg gggcgcacgc tgtgggaagc ggtacgccgg cacgcgcgcg 300 cgtacaacgc cacggtcata tggtacaaga tcgagagcgg gtgcgcccgg ccgctgtact 360 acatggagta caccgagtgc gagcccagga agcactttgg gtactgccgc taccgcacac 420 ccccgttttg ggacagcttc ctggcgggct tcgcctaccc cacggacgac gagctgggac 480 tgattatggc ggcgcccgcg cggctcgtcg agggccagta ccgacgcgcg ctgtacatcg 540 acggcacggt cgcctataca gatttcatgg tttcgctgcc ggccggggac tgctggttct 600 cgaaactcgg cgcggctcgc gggtacacct ttggcgcgtg cttcccggcc cgggattacg 660 ggttctgcgc agcaaaagaa tcacgcagta ctgacgtatc gaggcacaca ctacccgcag 720 aggccatagt cgactactgg ttcatgcgcc acgggggcgt cgttccgccg tattttgagg 780 agtcgaaggg ctacgagccg ccgcctgccg ccgatggggg ttcccccgcg ccacccggcg 840 acgacgaggc ccgcgagg at gaaggggaga ccgaggacgg ggcagccggg cgggagggca 900 acggcggccc cccaggaccc gaaggcgacg gcgagagtca gacccccgaa gccaacggag 960 cgagccgaaa gcgccgaggg cccggcccca gccccgacgc cgaccgcccc gaaggctggc 102 cgagcctcga agccatcacg caccccccgc ccgcccccgc tacgcccgct cgagctccgg 108 acgctgtttc ggtttctgtt ggtatcggta tcgctgctgc tgctatcgct tgcgttgctg 114 ctgctgctgc tggtgcttac ttcgtttata ttcgtcgtcg -ggtgctggt ccgctgccgc 120 aaaactgccg gtaaaccgaa gctttcggta acgttaacta ccgggttga cagtgctctg 125 < 210 > 17 < 211 > 418 < 212 > PRT < 213 > Bovine Herpesvirus 1 < 220 > < 221 > PEPTIDE < 222 > (1) .. (418) < 223 > sequence encoded by clone FlgD / Pots207nco (# 79) < 400 > 17 Met Glu Gly Pro Thr Leu Wing Val Leu Gly Wing Leu Leu Wing Val Wing 1 5 10 15 Val Ser Leu Pro Thr Pro Pro Wing Arg Val Thr Val Tyr Val Asp Pro 20 25 30 Pro Ala Tyr Pro Met Pro Arg Tyr Asn Tyr Thr Glu Arg Trp His Thr 35 40 45 Thr Gly Pro lie Pro Pro Phe Wing Asp Gly Arg Glu Gln Pro Val 50 55 60 Giu Val Arg Tyr Ala Thr Ser Ala Ala Ala Cys Asp Met Leu Ala Leu 65 70 75 80 lie Wing Asp Pro Gln Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg 85 90 95 His Wing Arg Wing Tyr Asn Wing Thr Val lie Trp Tyr Lys lie Glu Ser 100 105 110 Giy Cys Wing Arg Pro Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro 115 120 125 Arg Lys His Phe Gly Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp 130 135 140 Being Phe Leu Wing Gly Phe Wing Tyr Pro Thr Asp Asp Glu Leu Gly Leu 145 150 155 160 lie Met Ala Ala Pro Ala Arg Leu Val Glu Gly Gln Tyr Arg Arg Ala 165 170 175 Leu Tyr lie Asp Gly Thr Val Wing Tyr Thr Asp Phe Met Val Ser Leu 180 185 190 Pro Wing Gly Asp Cys Trp Phe Ser Lys Leu Gly Wing Wing Arg Gly Tyr 195 200 205 Thr Phe Gly Wing Cys Phe Pro Wing Arg Asp Tyr Glu Gln Lys Lys Val 210 215 220 Leu Arg Leu Thr Tyr Leu Thr Gln Tyr Tyr Pro Gln Glu Wing His Lys 225 230 235 240 Ala lie Val Asp Tyr Trp Phe Met Arg His Gly Gly Val Val Pro Pro 245 250 255? Yr Pr.e Giu Giu Ser Lys Gly Tyr Glu Pro Pro Pro Ala Wing Asp Gly 260 265 270 He and Ser Prc Wing Pro Pro Gly Asp Asp Glu Wing Arg Glu Asp Glu Gly 275 280 285 Glu Tr.r Giu Asp Gly Wing Wing Gly Arg Glu Giy Asn Gly Gly Pro Pro r33 295 300 Gly Pro Glu Gly Asp Gly Glu Ser Gln Thr Prc Glu Wing Asn Gly Gly 305 310 315 320 Wing Glu Giy Glu Pro Lys Pro Gly Pro Ser Prc Asp Wing Asp Arg Pro 325 330 335 Glu Gly Trp Pro Be Leu Glu Ala lie Thr His Pro Pro Pro Wing Pro 340 345 350 Ala Thr Pro Ala Arg Ala Pro Asp Ala Val Ser Val Ser Val Gly lie 355 360 365 Giy to Ala Ala Ala to Ala Ala Cys Ala Ala Ala Ala Ala Ala Gly 3 * "" "C 375 380 Ala Tyr Phe Val Tyr lie Arg Arg Arg Gly Ala Gly Pro Leu Pro Arg 385 390 395 400 Lys Pro Lys Lys Leu Pro Wing Phe Gly Asn Val Asn Tyr Ser Wing Leu 405 410 415 Pro Gly < 210 > 18 < 211 > 1405 < 212 > DNA < 213 > Bovine Herpesvirus 1 < 220 > < 221 > gene < 222 > (1) .. (1405) < 223 > gD of BHV-1 from M59846 Access to GenBank. < 40C > 18 gggccgcagc cccggctggg tatatatccc cgacgggcga ctagagatac actcgccccg 60 cgcggctgct gcgagcgggc gaacatgcaa gggccgacat tggccgtgct gggcgcgctg 120 ctcgccgttg cggtgagctt gcctacaccc gcgccgcggg tgacggtata cgtcgacccg 180 ccggcgtacc cgatgccgcg atacaactac actgaacgct ggcacactac cgggcccata 240 ccgtcgccct tcgcagacgg ccgcgagcag cccgtcgagg tgcgctacgc gacgagcgcg 300 gcsgcgtgcg acatgctggc gctgatcgca gacccgcagg tggggcgcac gctgtgggaa CGGT 360? gc ggcacgcgcg cgcgtacaac gccacggtca tatggtacaa gatcgagagc 420 gggtgcgccc ggccgctgta ctacatggag tacaccgagt gcgagcccag gaagcacttt 480 gggtactgcc gctaccgcac acccccgttc tgggacagct tcctggcggg cttcgcctac 540 cccacggacg acgagctggg actgattatg gcggcgcccg cgcggctcgt cgagggccag 600 taccgacgcg cgctgtacat cgacggcacg gtcgcctata cagatttcat ggtttcgctg 660 ccggccgggg actgctggtt ctcgaaactc ggcgcggctc gcgggtacac ctttggcgcg 720 tgcttcccgg cccgggatta cgagcaaaag aaggttctgc gcctgacgta tctcacgcag 780 tactacccgc aggaggcaca caaggccata gtcgactact ggttcatgcg ccacgggggc 840 gtcgttccgc cgtattttga ggagtcgaag ggctacgagc cgccgcctgc cgccgatggg 900 ggttcccccg cgccacccgg cgacgacgag gcccgcgagg atgaagggga gaccgaggac 960 ggggcagccg ggcgggaggg caacggcggc cccccaggac ccgaaggcga cggcgagagt 102 agacccccg aagccaacgg aggcgccgag ggcgagccga aacccggccc cagccccgac 108 gccgaccgcc ccgaaggctg gccgagcctc gaagccatca cgcacccccc gcccgccccc 114 gctacgcccg cggcccccga cgccgtgccg gtcagcgtcg ggatcggcat tgcggctgcg 120 gcgatcgcgt gcgtggccgc cgccgcc gcc ggcgcgtact tcgtctatac gcgccggcgc 126 ggtgcgggtc cgctgcccag aaagccaaaa aagctgccgg cctttggcaa cgtcaactac 132 agcgcgctgc ccgggtgagc ggcctaggcc ctcccccgac cqcccccttt gctcctagcc 138 ccggctcctg ccgagccgcg cgggg 140 < 210 > 19 < 211 > 417 < 212 > PRT < 213 > Bovine Herpesvirus 1 < 220 > < 221 > PEPTIDE < 222 > (1) .. (417) < 223 > gD of BHV-1 encoded by N ° M59846 Access to GenBank. < 400 > 19 Met Gln Gly Pro Thr Leu Wing Val Leu Gly Wing Leu Leu Wing Val Wing 1 5 10 15 Val Ser Leu Pro Thr Pro Pro Wing Arg Val Thr Val Tyr Val Asp Pro 20 25 30 Pro Ala Tyr Pro Met Pro Arg Tyr Asn Tyr Thr Glu Arg Trp His Thr 35 40 45 Thr Gly Pro lie Pro Pro Phe Wing Asp Gly Arg Glu Gln Pro Val 50 55 60 Glu Val Arg Tyr Ala Thr Ser Ala Ala Ala Cys Asp Met Leu Ala Leu 65 70 75 80 lie Wing Asp Pro Gln Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg 85 90 95 His Aia Arg Ala Tyr Asn Ala Thr Val lie Trp Tyr Lys lie Glu Ser 100 105 110 Gly Cys Aia Arg Pro Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro 115 120 125 Arg Lys Hi = Phe Gly Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp 130 135 140 Being Phe Leu Wing Gly Phe Wing Tyr Pro Thr Asp Asp Glu Leu Gly Leu 145 150 155 160 lie Met Ala Aia Pro Aia Arg Leu Val Glu Gly Gln Tyr Arg Arg Ala 165 170 175 Leu Tyr lie Asp Gly Thr Val Wing Tyr Thr Asp Phe Met Val Ser Leu 180 185 190 Pro Aia Giy Asp Cys Trp Phe Ser Lys Leu Gly Ala Wing Arg Gly Tyr 195 200 205 Thr Phe Gly Wing Cys Phe Pro Wing Arg Asp Tyr Glu Gln Lys Lys Val 210 215 220 Leu Arg Leu Thr Tyr Leu Thr Gln Tyr Tyr Pro Gln Glu Wing His Lys 225 230 235 240 Ala lie Val Asp Tyr Trp Phe Met Arg His Gly Gly Val Val Pro Pro 245 250 255 Tyr Phe Glu Glu Ser Lys Giy Tyr Glu Pro Pro Pro Wing Wing Asp Gly 260 265 270 Gly Pro Pro Wing Pro Pro Gly Asp Asp Glu Wing Arg Glu Asp Glu Gly 2"7? 280 285 Glu Thr Glu Asp Gly Wing Wing Gly Arg Glu Gly Asn Gly Gly Pro Pro 290 295 300 Gly Pro Glu Gly Asp Gly Glu Ser Gln Thr Pro Glu Wing Asn Gly Gly 305 310 315 320 Wing Glu Gly Glu Pro Lys Pro Gly Pro Ser Pro Asp Wing Asp Arg Pro 325 330 335 Glu Gly Trp Pro Ser Leu Glu Wing fie Thr His Pro Pro Pro Wing Pro 340 345 350 Ala Thr Pro Ala Ala Frc Asp Aia Val Pro Val Ser Val Gly lie Gly 355 360 365 lie Aia Ala Ala Ala lie Ala Cys Val Aia Ala Ala Ala Ala Ala Gly Ala 370 375 380 Tvr Phe Val Tyr Thr Arg Arg Arg Giy Wing Gly Pro Leu Pro Arg Lys 385 390 395 400 ? rc ..vs Lys Leu Pro A .a Pne Gly Asn Val Asn Tyr Ser Ala Leu Pro 405 41G 415 rz l < 210 > 20 < 211 > 1218 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: Sequence of pQE-tmgD that encodes a tmgD. < 400 > 20 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca 60 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga 120 atcaccatca ggatctcacc ccatacggat ccgcatgcca tgagcttgcc tacacccgcg 180 ccgcgggtga cggtatacgt cgacccgccg tgccgcgata gcgtacccga caactacact 240 gaacgctggc acactaccgg gcccataccg tcgcccttcg cagacggccg cgagcagccc 300 gtcgaggtgc gctacgcgac gagcgcggcg gcgtgcgaca tgctggcgct gatcgcagac 360 ccgcaggtgg ggcgcacgct gtgggaagcg gtacgccggc acgcgcgcgc gtacaacgcc 420 acggt atat ggtacaagat cgagagcggg tgcgcccggc cgctgtacta catggagtac 480 accgagtgcg agcccaggaa gcactttggg tactgccgct accgcacacc cccgttttgg 540 gacagcttcc tggcgggctt cgcctacccc acggacgacg agctgggact gattatggcg 60 gcgcccgcgc ggctcgtcga gggccagtac cgacgcgcgc tgtacatcga cggcacggtc 66 gcctatacag atttcatggt ttcgctgccg gccggggact gctggttctc gaaactcggc 72 gcggctcgcg ggtacacctt tggcgcgtgc ttcccggccc gggattacga gcaaaagaag 78 gttctgcgcc tgacgtatct cacgcagtac tacccgcagg aggcacacaa ggccatagtc 84 gactactggt tcatgcgcca cgggggcgtc gttccgccgt attttgagga gtcgaagggc 900 tacgagccgc cgcctgccgc cgatgggggt tcccccgcgc cacccggcga cgacgaggcc 96 cgcgasgatg aaggggagac cgaggacggg gcagccgggc gggagggcaa cggcggcccc 10 ccaggacccg aaggcgacgg cgagagtcag acccccgaag ccaacggagg cgccgagggc 108 < 210 > 21 < 211 > 367 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: tmgD encoded by pQE-tmgD. < 400 > 21 Met Arg Gly Ser His His His His His His His His Thr Asp Pro His Ala Met 1 5 10 15 Ser Leu Pro Thr Pro Pro Wing Arg Val Thr Val Tyr Val Asp Pro Pro 20 25 30 Wing Tyr Pro Met Pro Arg Tyr Asn Tyr Thr Glu Arg Trp His Thr Thr 40 45 Gly Pro lie Pro Pro Phe Wing Asp Gly Arg Glu Gln Pro Val Glu 50 55 60 Val Arg Tyr Ala Thr Ser Ala Ala Ala Cys Asp Met Leu Ala Leu lie 65 70 75 80 Wing Asp Pro Gln Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg His 35 90 95 Ala Arg Ala Tyr Asn Ala Thr Val lie Trp Tyr Lys lie Glu Ser Gly 100 105 110 Cys Ala Arg Pro Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro Arg 115 120 125 Lys His Phe Gly Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp Ser 130 135 140 Phe Leu Wing Gly Phe Wing Tyr Pro Thr Asp Asp Glu Leu Gly Leu lie 145 150 155 160 Met Wing Wing Pro Wing Arg Leu Val Glu Gly Gln Tyr Arg Arg Wing Leu 165 170 175 Tyr Lie Asp Giy Thr Val Wing Tyr Thr Asp Phe Met Val Ser Leu Pro 180 185 190 Wing Gly Asp Cys Trp Phe Ser Lys Leu Gly Wing Aia Arg Gly Tyr Thr 195 200 205 Phe Gly Wing Cys Phe Pro Wing Arg Asp Tyr Glu Gln Lys Lys Val Leu 210 215 220 Arg Leu Thr r Leu Thr Gin Tyr Tyr Pro Gln Giu Wing His Lys Wing 225 230 235 240 lie Val Asp Tyr Trp Phe Met Arg His Gly Giy Val Val Pro Pro Tyr 245 250 255 Phe Glu Glu Ser Lys Gly Tyr Glu Pro Pro Pro Wing Wing Asp Gly Gly 260 265 270 Ser Pro Pro Wing Pro Gly Asp Asp Glu Wing Arg Giu Asp Glu Gly Glu 275 280 285 Thr Glu Asp Gly Wing Wing Gly Arg Glu Gly Asn Gly Gly Pro Pro Gly 290 295 300 Pro Glu Gly Asp Gly Glu Ser Gln Thr Pro Glu Wing Asn Gly Gly Ala 305 310 315 320 Glu Gly Glu Pro Lys Pro Gly Pro Ser Pro Asp Wing Asp Arg Pro Glu 325 330 335 Giy Trp Pro Ser Leu Glu Ala lie Thr His Pro Pro Pro Ala Pro Wing 340 345 350 Thr Pro Ala Arg Ala Arg Tyr Pro Giy Ser Thr Cys Ser Gln Ala 355 360 365 < 210 > 22 < 211 > 1360 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: portion of pQE-GnRH: gD, including the sequence encoding 4GnRH-tmgD. < 400 > 22 ctcgagaaat cataaaaaat ttatttgct gtgagcgga taacaattat aatagattca 60 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga 120 atcaccatca ggatctcacc ccatacggat ccgcatgcca gcactggtca tggatccaga 180 tatggtctgc gtccgggtga acattggagc tacggtctac gccccgggga acactggtct 240 tatggcttac ggccgggaga gcattggagt tacggcctcc gtccaggttc catgagcttg 300 cctacacccg cgccgcgggt acggtatac gtcgacccgc cggcgtaccc gatgccgcga 360 tacaactaca ctgaacgctg gcacactac gggcccatac cgtcgccctt cgcagacggc 420 cgcgagcag ccgtcgaggt gcgctacgcg acgagcgcgg cggcgtgcga catgctggcg 480 ctgatcgcag acccgcaggt ggggcgcacg ctgtgggaag cggtacgccg gcacgcgcgc 540 gcgtacaacg ccacggtcat atggtacaag atcgagagcg ggtgcgcccg gccgctgtac 600 tacatggagt acaccgagtg cgagcccagg aagcactttg ggtactgccg ctaccgcaca 660 cccccgtttt gggacagctt cctggcgggc ttcgcctacc ccacggacga cgagctggga 720 ctgattatgg cggcgcccgc gcggctcgtc gagggccagt accgacgcgc gctgtacatc 780 gacggcacgg tcgcctatac agatttcatg gtttcgctgc cggccgggga ctgctggttc 840 tcgaaactcg gcgcggctcg cggg tacacc tttggcgcgt gcttcccggc ccgggattac 900 gagcaaaaga aggttctgcg cctgacgtat ctcacgcagt actacccgca ggaggcacac 960 aaggccatag tcgactactg gttcatgcgc cacgggggcg tcgttccgcc gtattttgag 102 gagtcgaagg gctacgagcc gccgcctgcc gccgatgggg gttcccccgc gccacccggc 108 gacgacgagg cccgcgagga tgaaggggag accgaggacg gggcagccgg gcgggagggc 114 aacggcggcc ccccaggacc cgaaggcgac ggcgagagtc agacccccga agccaacgga 120 ggcgccgagg gcgagccgaa acccggcccc agccccgacg ccgaccgccc cgaaggctgg 126 ccgagcctcg aagccatcac gcaccccccg cccgcccccg ctacgcccgc tcgagctcgg 132 taccccgggt cgacctgcag ccaagcttaa ttagctgagc 136 < 210 > 23 < 211 > 411 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: 4GnRH-tmgD encoded by pQE-GnRH: gD. < 400 > 23 Met Arg Gly Ser His His His His His His Hxs Thr Asp Pro His Ala Met 10 15 Asp Pro Glu His Trp Ser Tyr Gly Leu Arg Pro Gly Glu His Trp Ser 20 25 30 Tyr Gly Leu Arg Pro Gly Glu His rp Ser Tyr Gly Leu Arg Pro Gly 35 40 45 Glu His Trp Ser Tyr Gly Leu Arg Pro Gly Ser Met Ser Leu Pro Thr 50 55 60 Pro Ala Pro Arg Val Thr Val Tyr Val Asp Pro Pro Ala Tyr Pro Met 70 75 80 Pro Arg Tyr Asn Tyr Thr Glu Arg Trp His Thr Thr Gly Pro lie Pro 85 90 95 Ser Pro Phe Wing Asp Giy Arg Giu Gin Pro Val Giu Val Arg Tyr Wing 100 105 110 Thr Ser Ala Ala Aia Cys Asp Met Leu Ala Leu lie Ala Asp Pro Gln 115 120 125 Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg His Wing Arg Wing Tyr 130 135 140 Asn Wing Thr Val He Trp Tyr Lys He Glu Ser Gly Cys Wing Arg Pro 145 150 155 160 Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro Arg Lys His Phe Gly 165 170 175 Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp Ser Phe Leu Wing Gly 180 185 190 Phe Wing Tyr Pro Thr Asp Asp Glu Leu Gly Leu He Met Wing Wing Pro 195"200 205 Wing Arg Leu Val Glu Gly Gln Tyr Arg Arg Wing Leu Tyr He Asp Gly 210 215 220 Thr Val Ala Tyr Thr Asp Phe Met Val Ser Leu Pro Wing Gly Asp Cys 225 230 235 240 Trp Phe Ser Lys Leu Gly Wing Wing Arg Gly Tyr Thr Phe Gly Wing Cys 245 250 255 Phe Pro Wing Arg Asp Tyr Glu Gln Lys Lys Val Leu Arg Leu Thr Tyr 260 265 270 Leu Thr Gln Tyr Tyr Pro Gln Glu Wing His Lys Wing He Val Asp Tyr 275 280 285 Trp Phe Met Arg His Gly Gly Val Val Pro Pro Tyr Phe Glu Glu Ser 290 295 .300 Lys Gly Tyr Glu Pro Pro Pro Wing Wing Asp Gly Gly Ser Pro Wing Pro 305 310 315 320 Pro Gly Asp Asp Glu Wing Arg Glu Asp Glu Gly Glu Thr Glu Asp Gly 325 330 335 Ala Ala Gly Arg Glu Gly Asn Gly Gly Pro Pro Gly Pro Glu Gly Asp 340 345 350 Giy Glu Ser Gin Thr Pro Glu Wing Asn Gly Gly Wing Glu Gly Glu Pro 355 360 365 Lys Pro Gly Pro Ser Pro Asp Wing Asp Arg Pro Glu Gly Trp Pro Ser 370. 375 380 Leu Glu Wing He Thr His Pro Pro Pro Wing Pro Wing Thr Pro Wing Arg 385 390 395 400 Wing Arg Tyr Pro Gly Ser Thr Cys Ser Gln Wing 405 410 < 210 > 24 < 211 > 1360 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: portion of pQE-gD: GnRH, including the sequence encoding tmgD-4GnRH. < 400 > 24 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca 60 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga 12 atcaccatca ggatctcacc ccatacggat ccgcatgcca tgagcttgcc tacacccgcg 18 ccgcgggtga cggtatacgt cgacccgccg tgccgcgata gcgtacccga caactacact 24 gaacgctggc acactaccgg gcccataccg tcgcccttcg cagacggccg cgagcagccc 30 gtcgaggtgc gctacgcgac gagcgcggcg gcgtgcgaca tgctggcgct gatcgcagac 36 ccgcaggtgg ggcgcacgct gtgggaagcg gtacgccggc acgcgcgcgc gtacaacgcc 42 acggtcatat ggtacaagat cgagagcggg tgcgcccggc cgctgtacta catggagtac 48 accgagtgcg agcccaggaa gcactttggg tactgccgct accgcacacc cccgttttgg 54 gacagcttcc tggcgggctt cgcctacccc acggacgacg agctgggact gattatggcg 60 gcgcccgcgc ggctcgtcga gggccagtac cgacgcgcgc tgtacatcga cggcacggtc 66 gcctatacag atttcatggt ttcgctgccg gccggggact gctggttctc gaaactcggc 72 gcggctcgcg ggtacacctt tggcgcgtgc ttcccggccc gggattacga gcaaaagaag 78 gttctgcgcc tgacgtatct cacgcagtac tacccgcagg aggcacacaa ggccatagtc 84 gactactggt tcatgcgcca cgggggcg tc gttccgccgt attttgagga gtcgaagggc 90 tacgagccgc cgcctgccgc cgatgggggt tcccccgcgc cacccggcga cgacgaggcc 96 cgcgaggatg aaggggagac cgaggacggg gcagccgggc gggagggcaa cggcggcccc 10 ccaggacccg aaggcgacgg cgagagtcag acccccgaag ccaacggagg cgccgagggc 10 gagccgaaac ccggccccag ccccgacgcc gaccgccccg aaggctggcc gagcctcgaa 11 gccatcacgc accccccgcc cgcccccgct acgcccgctc gcactggtca gagctccaga 12 tatggtctgc gtccgggtga acattggagc tacggtctac gccccgggga acactggtct 12 tatggcttac ggccgggaga gcattggagt tacggcctcc gtccaggttg aagcttaatt 13 agctgagctt ggactcctgt tgatagatcc agtaatgacc 13 < 210 > 25 < 211 > 398 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: tmgD-4GnRH encoded by pQE-gD: GnRH. < 400 > 25 Met Arg Gly Ser His His His His His His His Thr Asp Pro His Ala Met 1 5 10 15 Ser Leu Pro Thr Pro Pro Wing Arg Val Thr Val Tyr Val Asp Pro Pro 20 25 30 Wing Tyr Pro Met Pro Arg Tyr Asn Tyr Thr Glu Arg Trp His Thr Thr 35 40 45 Giy Pro Pro Pro Pro Phe Wing Asp Giy Arg Glu Gln Pro Val Glu 50 55 60 Val Arg Tyr Ala Thr Ser Ala Ala Ala Cys Asp Met Leu Ala Leu He 65 70 75 80 Wing Asp Pro Gln Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg His 85 90 95 Ala Arg Ala Tyr Asn Ala Thr Val He Trp Tyr Lys He Glu Ser Gly 100 105 110 Cys Ala Arg Pro Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro Arg 115 120 125 Lys His Phe Gly Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp Ser 130 135 140 Phe Leu Wing Gly Phe Wing Tyr Pro Thr Asp Asp Glu Leu Gly Leu He 145 150 155 160 Met Wing Wing Pro Wing Arg Leu Val Glu Gly Gln Tyr Arg Arg Wing Leu 165 170 175 Tyr He Asp Giy Thr Val Wing Tyr Thr Asp Phe Met Val Ser Leu Pro 180 185 190 Wing Gly Asp Cys Trp Phe Ser Lys Leu Gly Wing Wing Arg Gly Tyr Thr 195 200 205 Phe Gly Aia Cys Phe Pro Wing Arg Asp Tyr Glu Gln Lys Lys Val Leu 210 215 220 : r Tyr iU yr Tyr Pro Gln Glu Ala His Lys Ala? C 235 240 He Val Asp Tyr Trp Phe Met Arg His Gly Gly Val Val Pro Pro Tyr 245 250 255 Phe Glu Glu Ser Lys Gly Tyr Glu Pro Pro Pro Wing Wing Asp Gly Gly 260 265 270 Ser Prc Aia Pro Pro Giy Asp Asp Glu Wing Arg Glu Asp Glu Gly Glu 275 280 285 Thr Glu Asp Gly Wing Wing Gly Arg Glu Gly Asn Gly Gly Pro Pro Gly 290 295 300 Pro Glu Giy Asp Gly Glu Be Gln Thr Pro Glu Wing Asn Gly Gly Ala 305 310 315 320 Glu Gly Glu Prc Lys Pro Gly Pro Ser Pro Asp Wing Asp Arg Pro Glu 325 330 335 Glv Ser Leu Glu Wing He Thr His Pro Pro Pro Wing Pro Wing 340 345 350 Thr Pro Wing Arg Wing Pro Glu His Trp Ser Tyr Gly Leu Arg Pro Gly 355 360 365 Glu His Trp Ser Tyr Gly Leu Arg Pro Gly Glu His Trp Ser Tyr Gly 370 375 380 Leu Arg Pro Gly Glu His Trp Ser Tyr Gly Leu Arg Pro Gly 385 390 395 < 210 > 26 < 21 1 > < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: portion of pQE-GnRH: gD: GnRH, including the sequence encoding 4GnRH-tmgD-4GnRH. < 400 > 26 ctcgagaaat catsasaaat ttatttgctt tgtgagcgga taacaattat aatagattca 60 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga 12 atcaccatca ggatctcacc ccatacggat ccgcatgcca gcactggtca tggatccaga 18 tatggtctgc gtccgggtga acattggagc tacggtctac gccccgggga acactggtct 24 tatggcttac ggccgggaga gcattggagt tacggcctcc gtccaggttc catgagcttg 30 cctacacccg cgccgcgggt gacggtatac gtcgacccgc cggcgtaccc gatgccgcga 36 tacaactaca ctgaacgctg gcacactacc gggcccatac cgtcgccctt cgcagacggc 42 gcgagcagc ccgtcgaggt gcgctacgcg acgagcgcgg cggcgtgcga catgctggcg 48 ctgatcgcag acccgcaggt ggggcgcacg ctgtgggaag cggtacgccg gcacgcgcgc 54 gcgtacaacg ccacggtcat atggtacaag atcgagagcg ggtgcgcccg gccgctgtac 60 tacatggagt acaccgagtg cgagcccagg aagcactttg ggtactgccg ctaccgcaca 66 cccccgtttt gggacagctt cctggcgggc ttcgcctacc ccacggacga cgagctggga 72 ctgattatgg cggcgcccgc gcggctcgtc gagggccagt accgacgcgc gctgtacatc 78 gacggcacgg tcgcctatac agatttcatg gtttcgctgc cggccgggga ctgctggttc 84 tcgaaactcg gcgcggctcg cgggtacacc tttggcgcgt gcttcccggc ccgggattac 90 gagcaaaaga aggttctgcg cctgacgtat ctcacgcagt actacccgca ggaggcacac 96 aaggccatag tcgactactg gttcatgcgc cacgggggcg tcgttccgcc gtattttgag 10 gagtcgaagg gctacgagcc gccgcctgcc gccgatgggg gttcccccgc gccacccggc 10 gacgacgagg cccgcgagga tgaaggggag accgaggacg gggcagccgg gcgggagggc 11 aacggcggcc ccccaggacc cgaaggcgac ggcgagagtc agacccccga agccaacgga 12 ggcgccgagg gcgagccgaa acccggcccc agccccgacg ccgaccgccc cgaaggctgg 12 ccgagcctcg aagccatcac gcaccccccg cccgcccccg ctacgcccgc tcgagctcca 13 gagcactggt catatggtct gcgtccgggt gaacattgga gctacggtct acgccccggg 13 gaacactggt cttatggctt acggccggga gagcattgga gttacggcct ccgtccaggt 14 14 < 210 > 27 < 211 > 442 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: 4-GnRH-tmgD-4GnRH encoded by pQE-GnRH: gD: GnRH. < 400 > 27 Met Arg Gly Ser His His His His His His His His Thr Asp Pro His Ala Met 1 5 10 15 Asp Pro Giu His Trp Ser Tyr Gly Leu Arg Pro Gly Glu His Trp Ser 20 25 30 Tyr Gly Leu Arg Pro Gly Glu His Trp Ser Tyr Gly Leu Arg Pro Gly 40 45 Glu His Trp Ser Tyr Giy Leu Arg Pro Giy Ser Met Ser Leu Pro Thr 50 55 60 Pro Ala Pro Arg Val Thr Val Tyr Val Asp Pro Pro Ala Tyr Pro Met 65 70 75 80 Pro Arg Tyr Asn Tyr Thr Glu Arg Trp His Thr Thr Gly Pro He Pro 85 90 95 Er -ro r-he Wing Asp Gly Arg Giu Gln Pro Val Glu Val Arg Tyr Ala 105 110 Thr Ser Wing Wing Wing Cys Asp Met Leu Wing Leu He Wing Asp Pro Gln 115 120 125 Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg His Wing Arg Wing Tyr 130 135 140 Asr. A. Thr Val lie Trp Tyr Lys He Giu Ser Gly Cys Aia Arg Pro 145 150 155 160 Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro Arg Lys His Phe Gly 165 170 175 Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp Ser Phe Leu Wing Gly 180 185 190 Phe Ala Tyr Pro Thr Asp Asp Glu Leu Gly Leu He Met Ala Ala Pro 195 200 205 Ala Arg Leu Val Glu Gly Gln Tyr Arg Arg Ala Leu Tyr He Asp Gly 210 215 220 Thr Val Ala Tyr Thr Asp Phe Met Val Ser Leu Pro Wing Gly Asp Cys 225 230 235 240 Trp Phe Ser Lys Leu Gly Wing Wing Arg Gly Tyr Thr Phe Gly Wing Cys 245 250 255 Phe Pro Wing Arg Asp Tyr Glu Gln Lys Lys Val Leu Arg Leu Thr Tyr 260 265 270 Leu Thr Gln Tyr Tyr Pro Gln Glu Wing His Lys Wing He Val Asp Tyr 275 280 285 Trp Phe Met Arg His Gly Gly Val Val Pro Pro Tyr Phe Glu Glu Ser 290 295 300 Lys Gly Tyr Giu Pro Pro Pro Wing Wing Asp Gly Giy Pro Pro Wing Pro 315 320 Pro Gly Asp Asp Glu Wing Arg Glu Asp Glu Gly Glu Thr Glu Asp Gly 325 330 335 Ala Ala Gly Arg Glu Gly Asn Gly Gly Pro Pro Giy Pro Glu Gly Asp 340 345 350 Giy Glu Ser Gin Thr Pro Glu Wing Asn Gly Gly Aia Glu Gly Glu Pro *. ' 360 365 Lys Pro Gly Pro Ser Pro Asp Wing Asp Arg Pro Glu Gly Trp Pro Ser 370 375 380 Leu Glu Wing He Thr His Pro Pro Pro Wing Pro Aia Thr Pro Wing Arg 385 390 395 400 Wing Pro Glu His Trp Ser Tyr Gly Leu Arg Pro Gly Glu His Trp Ser 405 410 415 Tyr Gly Leu Arg Pro Gly Glu His Trp Ser Tyr Gly Leu Arg Pro Gly 420 425 430 Glu His Trp Ser Tyr Gly Leu Arg Pro Gly 435 440 < 210 > 28 < 211 > 1079 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: portion of pCMV-tgD, including the sequence that encodes a truncated gD. < 400 > 28 gatatcatgc aggggccgac attggccgtg ctgggcgcgc tgctcgccgt tgcggtaagc 60 ttgcctacac ccgcgccgcg ggtgacggta tacgtcgacc cgccggcgta cccgatgccg 120 cgatacaact acactgaacg ctggcacact accgggccca taccgtcgcc cttcgcagac 180 ggccgcgagc agcccgtcga ggtgcgctac gcgacgagcg cggcggcgtg cgacatgctg 240 gcgctgatcg cagacccgca ggtggggcgc acgctgtggg aagcggtacg ccggcacgcg 300 cgcgcgtaca acgccacggt catatggtac aagatcgaga gcgggtgcgc ccggccgctg 360 tactacatg 5g c3ccg gtgcgagccc aggaagcact ttgggtactg ccgctaccgc 420 acacccccgt tttgggacag cttcctggcg ggcttcgcct accccacgga cgacgagctg 480 ggactgatta tggcggcgcc cgcgcggctc gtcgagggcc agtaccgacg cgcgctgtac 540 cggtcgccta atcgacggca tacagatttc atggtttcgc tgccggccgg ggactgctgg 600 tctcgaa c tcggcgcggc tcgcgggtac acctttggcg cgtgcttccc ggcccgggat 660 tacgagcaaa agaaggttct gcgcctgacg tatctcacgc agtactaccc gcaggaggca 720 tagtcgacta cacaaggcca ctggttcatg cgccacgggg gcgtcg cc gccgtatttt 780 agggctacga gaggagtcga gccgccgcct gccgccgatg ggggttcccc cgcgccaccc 840 ggcgacgacg aggcccgcga g gatgaaggg gagaccgagg acggggcagc cgggcgggag 900 ggcaacggcg gccccccagg acccgaaggc gacggcgaga gtcagacccc cgaagccaac 960 ggaggcgccg agggcgagcc gaaacccggc cccagccccg acgccgaccg ccccgaggct 102 ggccgagcct cgaagccatc acgcaccccc cgcccgcccc cgctacgccc tgaggtacc 107 < 210 > 29 < 211 > 353 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: truncated gD encoded by pCMV-tgD. < 400 > 29 Met Gln Gly Pro Thr Leu Ala Val Leu Gly Ala Leu Leu Ala Val Ser 1 5 10 15 Leu Pro Thr Pro Pro Wing Arg Val Thr Val Tyr Val Asp Pro Pro Wing 20 25 30 Tyr Pro Met Pro Arg Tyr Asn Tyr Thr Glu Arg Trp His Thr Thr Gly 40 45 Pro He Pro Ser Pro Phe Wing Asp Gly Arg Glu Gln Pro Val Glu Val 50 55 60 Arg Tyr Ala Thr Ser Ala Ala Ala Cys Asp Met Leu Ala Leu He Ala 65 70 75 80 Asp Pro Gln Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg Hxs Wing 85 90 95 Arg Ala Tyr Asn Ala Thr Val He Trp Tyr Lys He Glu Ser Gly Cys 100 105 110 Wing Arg Pro Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro Arg Lys 115 120 125 His Phe Giy Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp Ser Phe 130 135 140 .eu A_a ^ y r. Tyr Pro Thr Asp Asp Glu Leu Gly Leu He Met .45 150 155 160 Wing Wing Pro Wing Arg Leu Val Glu Giy Gln Tyr Arg Arg Wing Leu Tyr 165 170 175 He Asp Gly Thr Val Wing Tyr Thr Asp Phe Met Val Ser Leu Pro Wing 180 185 190 Gly Asp Cys Trp Phe Ser Lys Leu Gly Ala Wing Arg Gly Tyr Thr Phe 195 200 205 Giy Ala Cys Phe Pro Aia Arg Asp Tyr Glu Gln Lys Lys Val Leu Arg 2iC 215 220 Leu Thr Tyr Leu Thr Gln Tyr Tyr Pro Gln Glu Wing His Lys Ala He 225 230 235 240 Val Asp Tyr Trp Phe Met Arg His Gly Gly Val Val Pro Pro Tyr Phe 245 250 255 Glu Glu Ser Lys Gly Tyr Glu Pro Pro Pro Wing Wing Asp Gly Gly Ser 260 265 270 Pro Pro Wing Pro Giy Asp Asp Glu Aia Arg Glu Asp Glu Gly Glu Thr 275 280 285 Glu Asp Gly Ala Ala Gly Arg Glu Gly Asn Gly Gly Pro Pro Gly Pro 290 295 300 Glu Gly Asp Gly Glu Ser Gln Thr Pro Glu Wing Asn Gly Gly Ala Glu 305 310 315 320 Gly Glu Pro Lys Pro Gly Pro Ser Pro Asp Wing Asp Arg Pro Glu Gly 325 330 335 Trp Pro Ser Leu Glu Wing He Thr His Pro Pro Pro Wing Pro Thr Wing 340 345 350 Pro < 210 > 30 < 211 > 1241 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: portion of pCMV-gD: GnRH, including the sequence encoding a fusion protein of tgD-4GnRH. < 400 > 30 gcggccgcaa gatatcatgc aggggccgac attggccgtg ctgggcgcgc tgctcgccgt 60 tgcggtaagc ttgcctacac ccgcgccgcg ggtgacggta tacgtcgacc cgccggcgta 12 cccgatgccg cgatacaact acactgaacg ctggcacact accgggccca taccgtcgcc 18 cttcgcagac ggccgcgagc agcccgtcga ggtgcgctac gcgacgagcg cggcggcgtg 24 cgacatgctg gcgctgatcg cagacccgca ggtggggcgc acgctgtggg aagcggtacg 30 ccggcacgcg cgcgcgtaca acgccacggt catatggtac aagatcgaga gcgggtgcgc 36 ccggccgctg tactacatgg agtacaccga gtgcgagccc aggaagcact ttgggtactg 42 ccgctaccgc acacccccgt tttgggacag cttcctggcg ggcttcgcct accccacgga 48 cgacgagctg ggactgatta tggcggcgcc cgcgcggctc gtcgagggcc agtaccgacg 54 atcgacggca cgcgctgtac cggtcgccta tacagatttc atggtttcgc tgccggccgg 60 ggactgctgg ttctcgaaac tcggcgcggc tcgcgggtac acctttggcg cgtgcttccc 66 ggcccgggat tacgagcaaa agaaggttct gcgcctgacg tatctcacgc agtactaccc 72 cacaaggcca gcaggaggca tagtcgacta ctggttcatg cgccacgggg gcgtcgttcc 78 gaggagtcga gccgtatttt agggctacga gccgccgcct gccgccgatg ggggttcccc 84 cgcgccaccc ggcgacgacg aggcccgcg to ggatgaaggg gagaccgagg acggggcagc 90 cgggcgggag ggcaacggcg gccccccagg acccgaaggc gacggcgaga gtcagacccc 96 cgaagccaac ggaggcgccg agggcgagcc gaaacccggc cccagccccg acgccgaccg 10 ccccgaaggc tggccgagcc tcgaagccat cacgcacccc ccgcccgccc ccgctacgcc 10 cgctcgagct ccagagcact ggtcatatgg tctgcgtccg ggtgaacatt ggagctacgg 11 tctacgcccc ggggaacact ggtcttatgg cttacggccg ggagagcatt ggagttacgg 12 cctccgtcca ggttgaagct gggatactag tgagcggccg c 12 < 210 > 31 < 211 > 397 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: tgD-4GnRH fusion protein encoded by pCMV-gD: GnRH. < 400 > 31 Met Gln Gly Pro Thr Leu Ala Val Leu Gly Ala Leu Leu Ala Val Ser 1 5 10 15 Leu Pro Thr Pro Pro Wing Arg Val Thr Val Tyr Val Asp Pro Pro Wing 20 25 30 Tyr Pro Met Pro Arg Tyr Asn Tyr Thr Glu Arg 7rp His Thr Thr Gly 35 '40 45 Prc Lie Pro Ser Pro Phe Wing Asp Giy Arg Glu Gln Pro Val Glu Val 50 55 60 Arg Tyr Ala Thr Ser Ala Ala Ala Cys Asp Met Leu Ala Leu He Ala 65 70 75 80 Asp Pro Gln Val Giy Arg Thr Leu Trp Glu Wing Val Arg Arg His Wing 85 90 95 Arg Aia Tyr Asr. Wing Thr Val He Trp Tyr Lys He Glu Ser Gly Cys 100 105 110 Wing Arg Pro Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro Arg Lys 115 120 125 His Phe Giy Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp Ser Phe 130 135 140 Leu Ala Gly phe Aia Tyr Pro Thr Asp Asp Glu Leu Gly Leu He Met 145 150 155 160 Ala Ala Pro Aia Arg Leu Val Glu Gly Gln Tyr Arg Arg Ala Leu Tyr 165 170 175 He Asp Gly Thr Val Wing Tyr Thr Asp Phe Met Val Ser Leu Pro Wing 180 185 190 Gly Asp Cys Trp Phe Ser Lys Leu Gly Ala Wing Arg Gly Tyr Thr Phe 195 200 _ 205 Gly Aia Cys Phe Pro Wing Arg Asp Tyr Glu Gln Lys Lys Val Leu Arg 210 215 220 Leu Thr Tyr Leu Thr Gln Tyr Tyr Pro Gln Glu Wing His Lys Ala lie 225 • 230 235 240 Val Asp Tyr Trp Phe Met Arg His Gly Gly Val Val Pro Pro Tyr Phe 245 250 255 Glu Glu Gly Ser Gly Tyr Glu Pro Pro Wing Gly Asp Gly Gly Ser 260 265 270 Pro Pro Wing Gly Asp Asp Glu Wing Arg Glu Asp Glu Glu Glu Thr 275 280 285 Glu Asp Giy Wing Wing Giy Arg Giu Giy Asn Gly Gly Pro Pro Gly Pro 290 295 300 Glu Gly Asp Gly Glu Ser Gln Thr Pro Glu Wing Asn Gly Gly Ala Glu 305 310 315 320 Gly Glu Pro Lys Pro Giy Pro Ser Pro Asp Wing Asp Arg Pro Glu Gly 325 330 335 Trp Pro Ser Leu Glu Aia He Thr His Pro Pro Pro Ala Pro Wing Thr 340 345 350 Pro .Ala Arg Ala Pro Glu His Trp Ser Tyr Gly Leu Arg Pro Gly Glu 355 360 365 His Trp Ser Tyr Gly Leu Arg Pro Gly Glu His Trp Ser Tyr Gly Leu 370 375 380 Arg Pro Giy Glu His Trp Ser Tyr Gly Leu Arg Pro Gly 385 390 395 < 210 > 32 < 211 > 120 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: sequence that encodes a GnRH tetramer. < 400 > 32 gagcactggt catatggtct gcgtccgggt gaacattgga gctacggtct acgccccggg 60 gaacactggt cttatggctt acggccggga gagcattgga gttacggcct ccgtccaggt 120 < 210 > 33 < 211 > 30 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: sequence that encodes a GnRH monomer. < 400 > 33 gagcactggt catatggtct gcgtccgggt 30 < 210 > 34 < 211 > 1179 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: sequence that encodes a fusion protein of 4GnRH-tmgD. < 400 > 34 gagcactggt catatggtct gcgtccgggt gaacattgga gctacggtct acgccccggg 60 gaacactggt cttatggctt acggccggga gagcattgga gttacggcct ccgtccaggt 12 t catgagct tgcctacacc cgcgccgcgg gtgacggtat acgtcgaccc gccggcgtac 18 gatacaacta ccgatgccgc caccgaacgc tggcacacta ccgggcccat accgtcgccc 24 ttcgcagacg gccgcgagca gcccgtcgag gtgcgctacg cgacgagcgc ggcggcgtgc 30 gacatgctgg cgctgatcg agacccgcag g-cggggcgca cgctgtggga agcggtacgc 36 gcgcgtacaa cggcacgcgc cgccacggtc atatggtaca agatcgagag cgggtgcgcc 42 cggccgctgt actacatgga gtacaccgag tgcgagccca ggaagcactt tgggtactgc 48 cgctaccgca cacccccgtt ttgggacagc ttcctggcgg gcttcgccta ccccacggac 54 gacgagctgg gactgattat ggcggcgccc gcgcggctcg tcgagggcca gtaccgacgc 60 gcgctgtaca tcgacggcac ggtcgcctat acagatttca tggtttcgct gccggccggg 66 gactgctggt tctcgaaact cggcgcggct cgcgggtaca cctttggcgc gtgcttcccg 72 gcccgggatt acgagcaaaa gaaggttctg cgcctgacgt atctcacgca gtactacccg 78 caggaggcac acaaggccat agtcgactac tggttcatgc gccacggggg cgtcgttccg 84 ccgtattttg aggagtcgaa gggctacga g ccgccgcctg ccgccgatgg gggttccccc 90 gcgccacc g gcgacgacga ggcccgcgag gatgaagggg agaccgagga cggggcagcc 96 gggcgggagg gcaacggcgg ccccccagga cccgaaggcg acggcgagag tcagaccccc 10 gaagccaacg gaggcgccga gggcgagccg aaacccggcc ccagccccga cgccgaccgc 10 cccgaaggct ggccgagcct cgaagccatc acgcaccccc cgcccgcccc cgctacgccc 11 gctcgagctc ggtaccccgg gtcgacctgc agccaagct 11 < 210 > 35 < 211 > 340 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: a truncated mature gD of BHV-1. 30 Pro He Pro Ser Pro Phe Wing Asp Gly Arg Glu Gln Pro Val Glu Val 35 40 45 Arg Tyr Ala Thr Ser Ala Aia Aia Cys Asp Met Leu Ala Leu He Ala 5C 35 60 Asp Pro Gln Val Gly Arg Thr Leu Trp Glu Wing Val Arg Arg His Wing 65 70 75 80 Arg Ala Tyr Asn Ala Thr Val He Trp Tyr Lys He Glu Ser Gly Cys 85 90 95 Wing Arg Pro Leu Tyr Tyr Met Glu Tyr Thr Glu Cys Glu Pro Arg Lys 1JG 105 110 His Phe Gly Tyr Cys Arg Tyr Arg Thr Pro Pro Phe Trp Asp Ser Phe 115 120 125 Leu Wing Gly Phe Wing Tyr Pro Thr Asp Asp Glu Leu Gly Leu He Met 130 135 140 Wing Wing Pro Wing Arg Leu Val Glu Gly Gln Tyr Arg Arg Ala Leu Tyr 145 150 155 160 He Asp Gly Thr Val Wing Tyr Thr Asp Phe Met Val Ser Leu Pro Wing 165 170 175 Gly Asp Cys Trp Phe Ser Lys Leu Gly Ala Wing Arg Gly Tyr Thr Phe 180 185 190 Gly Ala Cys Phe Pro Ala Arg Asp Tyr Glu Gln Lys Lys Val Leu Arg 195 200 205 Leu Thr Tyr Leu Thr Gln Tyr Tyr Pro Gln Glu Wing His Lys Wing He 210 215 220 Val Asp Tyr Trp Phe Met Arg His Gly Gly Val Val Pro Pro Tyr Phe 225 230 235 240 Glu Glu Ser Lys Gly Tyr Glu Pro Pro Pro Wing Wing Asp Gly Gly Ser 245 250 255 Pro Wing Pro Pro Gly Asp Asp Glu Wing Arg Glu Asp Glu Gly Glu Thr 260 265 270 Glu Asp Gly Ala Ala Gly Arg Glu Gly Asn Gly Gly Pro Pro Gly Pro 275 280 285 Glu Gly Asp Gly Glu Ser Gln Thr Pro Glu Wing Asn Gly Gly Ala Glu 290 295 300 Gly Glu Pro Lys Pro Gly Pro Ser Pro Asp Wing Asp Arg Pro Glu Gly 305 310 315 320 Trp Pro Ser Leu Glu Wing He Thr His Pro Pro Pro Wing Pro Thr Wing 325 330 335 Pro Ala Arg Wing 340 < 210 > 36 < 211 > 1020 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: sequence that encodes a truncated mature gD of BHV-1. < 400 > 36 ttgcctacac ccgcgccgcg ggtgacggta tacgtcgacc cgccggcgta cccgatgccg 60 cgatacaact acactgaacg ctggcacact accgggccca taccgtcgcc cttcgcagac 120 ggccgcgagc agcccgtcga ggtgcgctac gcgacgagcg cggcggcgtg cgacatgctg 180 gcgctgatcg cagacccgca ggtggggcgc acgctgtggg aagcggtacg ccggcacgcg 240 cgcgcgtaca acgccacggt catatggtac aagatcgaga gcgggtgcgc ccggccgctg 300 tactacatgg agtacaccga gtgcgagccc aggaagcact ttgggtactg ccgctaccgc 360 acacccccgt tttgggacag cttcctggcg ggcttcgcct accccacgga cgacgagctg 420 ggactgatta tggcggcgcc cgcgcggctc gtcgagggcc agtaccgacg cgcgctgtac 480 cggtcgccta atcgacggca tacagatttc atggtttcgc tgccggccgg ggactgctgg 540 ttctcgaaac tcggcgcggc tcgcgggtac acctttggcg cgtgcttccc ggcccgggat 600 tacgagcaaa agaaggttct gcgcctgacg tatctcacgc agtactaccc gcaggaggca 660 tagtcgacta cacaaggcca ctggttcatg cgccacgggg gcgtcgttcc gccgtatttt 720 agggctacga gaggagtcga gccgccgcct gccgccgatg ggggttcccc cgcgccaccc 780 ggcgacgacg aggcccgcga ggatgaaggg gagaccgagg acggggcagc cgggcgggag 840 ggcaacggcg gcccccc agg acccgaaggc gacggcgaga gtcagacccc cgaagccaac 900 ggaggcgccg agggcgagcc gaaacccggc cccagccccg acgccgaccg ccccgaaggc 960 tggccgagcc tcgaagccat cacgcacccc ccgcccgccc ccgctacgcc cgctcgagct 102 < 210 > 37 < 211 > 15 < 212 > PRT < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: leader of 6XHIS. < 40C > 37 Met Arg Giy Ser His His His His His His His His Thr Asp Pro His Ala 1 5 10 15 < 210 > 38 < 211 > 45 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: sequence that encodes the leader of 6XHIS. < 400 > 38 atgagaggat ctcaccatca ccatcaccat acggatccgc atgcc 45 < 210 > 39 < 211 > 1017 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: open reading frame for the 6XHIS leader, the truncated mature gD, and the GnRH tetramer encoded by bac-gD: GnRH. < 400 > 39 atgagcttgc ctacacccgc gccgcgggtg acggtatacg tcgacccgcc ggcgtacccg 60 atgccgcgat acaactacac tgaacgctgg cacactaccg ggcccatacc gtcgcccttc 120 gcagacggcc gcgagcagcc cgtcgaggtg cgctacgcga cgagcgcggc ggcgtgcgac 180 atgctggcgc tgatcgcaga cccgcaggtg gggcgcacgc tgtgggaagc gg acgccgg 240 cacgcgcgcg cgtacaacgc cacggtcata tggtacaaga tcgagagcgg gtgcgcccgg 300 ccgctgtact acatggagta caccgagtgc gagcccagga agcactttgg gtactgccgc 360 taccgcacac ccccgttttg ggacagcttc ctggcgggct tcgcctaccc cacggacgac 420 gagctgggac tgattatggc ggcgcccgcg cggctcgtcg agggccagta ccgacgcgcg 480 ctgtacatcg acggcacggt cgcctataca gatttcatgg tttcgctgcc ggccggggac 540 tgctggttct cgaaactcgg cgcggctcgc gggtacacct ttggcgcgtg cttcccggcc 600 cgggattacg agcaaaagaa ggttctgcgc ctgacgtatc tcacgcagta ctacccgcag 660 gaggcacaca aggccatagt cgactactgg ttcatgcgcc acgggggcgt cgttccgccg 720 tattttgagg agtcgaaggg ctacgagccg ccgcctgccg ccgatggggg ttcccccgcg 780 ccacccggcg acgacgaggc ccgcgaggat gaaggggaga ccgaggacgg ggcagccggg 840 cgggagggca acggcg gccc cccaggaccc gaaggcgacg gcgagagtca gacccccgaa 900 gccaacggag gcgccgaggg cgagccgaaa cccggcccca gccccgacgc cgaccgcccc 96 gaaggctggc cgagcctcga agccatcacg caccccccgc ccgcccccgc tacgccc 10 < 210 > 40 < 21 1 > 1272 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: sequence that encodes a fusion protein of 4GnRH-tmgD-4GnRH. < 400 > 40 gagcactggt catatggtct gcgtccgggt gaacattgga gctacggtct acgccccggg 60 gaacactggt cttatggctt acggccggga gagcattgga gttacggcct ccgtccaggt 120 tccatgagct tgcctacacc cgcgccgcgg gtgacggtat acgtcgaccc gccggcgtac 180 ccgatgccgc gatacaacta cactgaacgc tggcacacta ccgggcccat accgtcgccc 240 ttcgcagacg gccgcgagca gcccgtcgag gtgcgctacg cgacgagcgc ggcggcgtgc 300 gacatgctgg cgctgatcgc agacccgcag gtggggcgca cgctgtggga agcggtacgc 360 gcgcgtacaa cggcacgcgc cgccacggtc atatggtaca agatcgagag cgggtgcgcc 420 cggccgctgt actacatgga gtacaccgag tgcgagccca ggaagcactt tgggtactgc 480 cgctaccgca cacccccgtt ttgggacagc ttcctggcgg gcttcgccta ccccacggac 540 gacgagctgg gactgattat ggcggcgccc gcgcggctcg tcgagggcca gtaccgacgc 600 gcgctgtaca tcgacggcac ggtcgcctat acagatttca tggtttcgct gccggccggg 660 gactgctggt tctcgaaact cggcgcggct cgcgggtaca cctttggcgc gtgcttcccg 720 gcccgggatt acgagcaaaa gaaggttctg cgcctgacgt atctcacgca gtactacccg 780 caggaggcac acaaggccat agtcgactac tggttcatgc gccacggggg cgtcgttccg 840 ccgtattttg aggagtc gaa gggctacgag ccgccgcctg ccgccgatgg gggttccccc 900 gcgccacccg gcgacgacga ggcccgcgag gatgaagggg agaccgagga cggggcagcc 960 gggcgggagg gcaacggcgg ccccccagga cccgaaggcg acggcgagag tcagaccccc 102 gaagccaacg gaggcgccga gggcgagccg aaacccggcc ccagccccga cgccgaccgc 108 cccgaaggct ggccgagcct cgaagccatc acgcaccccc cgcccgcccc cgctacgccc 114 gctcgagctc cagagcactg gtcatatggt ctgcgtccgg gtgaacattg gagctacggt 120 ctacgccccg gggaacactg gtcttatggc ttacggccgg gagagcattg gagttacggc 126 ctccgtccag gt 127 < 210 > 41 < 211 > 1144 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: sequence that encodes a fusion protein of tmgD-4GnRH < 400 > 41 cttgcctaca cccgcgccgc gggtgacggt atacgtcgac ccgccggcgt acccgatgcc 60 gcgatacaac tacactgaac gctggcacac taccgggccc ataccgtcgc ccttcgcaga 120 cggccgcgag cagcccgtcg aggtgcgcta cgcgacgagc gcggcggcgt gcgacatgct 180 ggcgctgatc gcagacccgc aggtggggcg cacgctgtgg gaagcggtac gccggcacgc 240 gcgcgcgtac aacgccacgg tcatatggta caagatcgag agcgggtgcg cccggccgct 300 gtactacatg gagtacaccg agtgcgagcc caggaagcac tttgggtact gccgctaccg 360 cacacccccg t ttgggaca gcttcctggc gggcttcgcc taccccacgg acgacgagct 420 gggactgatt atggcggcgc ccgcgcggct cgtcgagggc agtaccgac gcgcgctgta 480 catcgacggc acggtcgcct atacagattt catggtttcg ctgccggccg gggactgctg 540 gttctcgaaa ctcggcgcgg ctcgcgggta cacctttggc gcgtgcttcc cggcccggga 600 ttacgagcaa aagaaggttc tgcgcctgac gtatctcacg cagtactacc cgcaggaggc 660 acacaaggcc atagtcgact actggttcat gcgccacggg ggcgtcgttc cgccgtattt 720 tgaggagtcg aagggctacg agccgccgcc tgccgccgat gggggttccc ccgcgccacc 780 cggcgacgac gaggcccgcg aggatgaagg ggagaccgag gacggggcag ccgggcggga 840 gggcaacggc ggcccccc ag gacccgaagg cgacggcgag agtcagaccc ccgaagccaa 900 cggaggcgcc gagggcgagc cgaaacccgg ccccagcccc gacgccgacc gccccgaagg 960 ctggccgagc ctcgaagcca tcacgcaccc cccgcccgcc cccgctacgc 102 ccgctcgagc tccagagcac tggtcatatg gtctgcgtcc gggtgaacat tggagctacg gtctacgccc 108 cggggaacac tggtcttatg gcttacggcc gggagagcat tggagttacg gcctccgtcc AGGT 114 114 < 210 > 42 < 211 > 23 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: primer P14-S1. < 400 > 42 ggagctccag agcactggtc ata 23 < 210 > 43 < 211 > 24 < 212 > DNA < 213 > Artificial sequence < 220 > < 223 > Description of the Artificial Sequence: primer P14-A138 < 400 > 43 aaagcttcaa cctggacgga ggcc 24 < 210 > 44 < 211 > 215 < 212 > PRT < 213 > Actinobacillus pleuropneumoniae < 400 > 44 Met Lys Lys Aia Val Leu Ala Wing Val Leu Gly Gly Ala Leu Leu Ala i 5 10 15 Gly Being Wing Met Wing His Gln Wing Gly Asp Val He Phe Arg Wing Gly 20 25 30 Wing He Gly Val He Wing Asn Being Ser Asp Tyr Gln Thr Gly Wing 35 40 45 Asp Val Asn Leu Asp Val Asn Asn Asn He Gln Leu Gly Leu Thr Gly 50 55 60 Thr Tyr Met Leu Ser Asp Asn Leu Gly Leu Glu Leu Leu Ala Wing Thr 65 70 75 80 Pro Phe Ser His Lys He Thr Gly Lys Leu Gly Wing Thr Asp Leu Gly 85 90 95 Glu Val Wing Lys Val Lys His Leu Pro Pro Ser Leu Tyr Leu Gln Tyr ICC 105 110 Tyr Phe Phe Asp Ser Asn Wing Thr Val Arg Pro Tyr Val Gly Wing Gly 115 120 125 Leu Asn Tyr Thr Arg Phe Phe Ser Wing Glu Ser Leu Lys Pro Gln Leu 130 135 140 Val Gln Asn Leu Arg Val Lys Lys His Ser Val Ala Pro He Ala Asn 145 150 155 160 Leu Gly Val Asp Val Lys Leu Thr Asp Asn Leu Ser Phe Asn Ala Wing 165 170 175 Wing Trp Tyr Thr Arg He Lys Thr Thr Wing Asp Tyr Asp Val Pro Gly 180 185 190 Leu Gly His Val Ser Thr Pro He Thr Leu Asp Pro Val Val Leu Phe 195 200 205 Ser Gly He Ser Tyr Lys Phe 210 215 < 210 > 45 < 211 > 364 < 212 > PRT < 213 > Actinobacillus pleuropneumoniae < 400 > 45 Met Lys Lys Ser Leu Val Ala Leu Thr Val Leu Ser Ala Ala Ala Val 1 5 10 15 Wing Gln Wing Aia Pro Gln Gln Asn Thr Phe Tyr Wing Gly Wing Lys Wing 20 25 30 Giy Trp Aia Ser Phe His Asp Gly He Glu Gln Leu Asp Ser Ala Lys 35 40 45 Asn Thr Asp Arg Gly Thr Lys Tyr Gly He Asn Arg Asn Ser Val Thr 50 55 60 Tyr Gly Val Phe Gly Gly Tyr Gln He Leu Asn Gln Asp Lys Leu Gly 65 70 75 80 Leu Ala Aia Glu Leu Gly Tyr Asp Tyr Phe Gly Arg Val Arg Gly Ser 85 90 95 Glu Lys Pro Asn Gly Lys Wing Asp Lys Lys Thr Phe Arg His Wing Ala 100 105 110 His Gly Ala Thr He Ala Leu Lys Pro Ser Tyr Glu Val Leu Pro Asp 115 120 125 Leu Asp Val Tyr Gly Lys Val Gly lie Wing Leu Val Asn Asn Thr Tyr 130 135 140 Lys Thr Phe Asn Wing Wing Gln Glu Lys Val Lys Thr Arg Arg Phe Gln 145 150 155 160 Being Ser Leu He Leu Gly Ala Gly Val Glu Tyr Ala He Leu Pro Glu 165 170 175 Leu Ala Ala Arg Val Glu Tyr Gln Trp Leu Asn Asn Wing Gly Lys Wing 180 185 190 Ser Tyr Ser Thr Leu Asn Arg Met Gly Wing Thr Asp Tyr Arg Ser Asp 195 200 205 lie Be Ser Val Be Wing Gly Leu Be Tyr Arg Phe Gly Gln Gly Wing 210 215 220 Val Pro Val Aia Aia Pro Wing Val Giu Thr Lys Asn Phe Wing Phe Ser 225 230 235 240 Be Asp Val Leu Phe Wing Phe Gly Lys Ser Asn Leu Lys Pro Wing Wing 245 250 255 Ala Thr Ala Leu Asp Ala Met Gln Thr Glu He Asn Asn Ala Gly Leu 260 265 270 Being Asn Ala Ala He Gin Val Asn Gly Tyr Thr Asp Arg He Gly Lys 275 280 285 Glu Aia Ser Asn Leu Lys Leu Ser Gln Arg Arg Ala Glu Thr Val Ala 290 295 300 Asn Tyr He Val Ser Lys Gly Ala Pro Ala Ala Asn Val Thr Ala Val 305 310 315 320 Gly Tyr Gly Glu Wing Asn Pro Val Thr Gly Ala Thr Cys Asp Lys Val 325 330 335 Lys Gly Arg Lys Wing Leu He Wing Cys Leu Wing Pro Asp Arg Arg Val 340 345 350 Glu Val Gln Val Gln Gly Thr Lys Glu Val Thr Met 355 360 < 210 > 46 < 211 > 369 < 212 > PRT < 213 > Actinobacillus pleuropneumoniae < 400 > 46 Met Lys Lys Ser Leu Val Ala Leu Ala Val Leu Ser Ala Ala Ala Ala 1 5 10 15 Ala Gln Ala Ala Pro Gln Gln Asn Thr Phe Tyr Ala Gly Ala Lys Val 20 25 30 Gly Gln Ser Ser Phe His His Gly Val Asn Gln Leu Lys Ser Gly His 40 45 Asp Asp Arg Tyr Asn Asp Lys Thr Arg Lys Tyr Gly lie Asn Arg Asn 50 55 60 Ser Val Thr Tyr Gly Val Phe Gly Gly Tyr Gln He Leu Asn Gln Asn 65 70 75 80 Asn Phe Gly Leu Wing Thr Glu Leu Gly Tyr Asp Tyr Tyr Gly Arg Val 85 90 95 Arg Gly Asn Asp Gly Glu Phe Arg Wing Met Lys His Ser Wing His Gly 100 105 110 Leu A-sn Phe Aia Leu Lys Pro Ser Tyr Giu Val Leu Pro Asp Leu Asp 115 120 125 Val Tyr Gly Lys Val Gly Val Val Wing Val Arg Asn Asp Tyr Lys Ser 130 135 140 Tyr Gly Ala Giu Asn Thr Asn Glu Pro Thr Glu Lys Phe His Lys Leu 145 150 155 160"Lys Wing Being Thr He Leu Gly Wing Gly Val Glu Tyr Wing He Leu Pro 165 170 175 Glu Leu Wing Aia Arg Val Glu Tyr Gln Tyr Leu Asn Lys Wing Gly Asn 180 185 190 Leu Asn Lys Ala Leu Val Arg Ser Gly Thr Gln Asp Val Asp Phe Gln 195 200 205 Tyr Ala Pro Asp He His Ser Val Thr Ala Gly Leu Ser Tyr Arg Phe 210 215 220 Gly Gln Gly Ala Val Ala Pro Val Val Glu Pro Glu Val Val Thr Lys 225 230 235 240 Asn Phe Wing Phe Ser Being Asp Val Leu Phe Asp Phe Gly Lys Ser Ser 245 250 255 Leu Lys Pro Ala Ala Ala Thr Ala Leu Asp Ala Ala Asn Thr Glu He 260 265 270 Wing Asn Leu Gly Leu Wing Thr Pro Wing He Gln Val Asn Gly Tyr Thr 275 280 285 Asp Arg He Gly Lys Glu Wing Being Asn Leu Lys Leu Being Gln Arg Arg 290 295 300 Wing Glu Thr Val Wing Asn Tyr Leu Val Ser Lys Gly Gln Asn Pro Wing 305 310 315 320 Asn Val Thr Wing Val Gly Tyr Gly Glu Wing Asn Pro Val Thr Gly Wing 325 330 335

Claims (17)

NOVELTY OF THE INVENTION CLAIMS

1. - A fusion protein to produce a double immune response in a vertebrate, whose fusion protein comprises: (a) a first protein portion analogous to all or a part of a peptide endogenously synthesized within the vertebrate, the activity of which peptide it must be inhibited within the vertebrate, and whose protein portion is itself incapable of producing an effective immune-suppressive response in said vertebrate, connected with (b) a second protein portion analogous to all or a part of an immunogen from of a pathogen, whose pathogen is capable of infecting the vertebrate pathogenically; causing portion (b) that the immune system of the vertebrate recognizes the portion (a) and produces a response that: (i) inhibits the activity of the peptide synthesized endogenously within the vertebrate; and (ii) protect the vertebrate with respect to an infection by the pathogen, when the vertebrate is vaccinated with an effective amount of the fusion protein.

2. A fusion protein according to claim 1, comprising a portion (a) analogous to all or a part of a GnRH peptide and a portion (b) analogous to all or a portion of an antigen of the BHV-1.

3. A fusion protein for producing an immune response in a vertebrate, whose fusion protein comprises: (a) a first protein portion analogous to all or a portion of a peptide whose activity is to be inhibited within the vertebrate, and whose protein portion is itself incapable of producing an effective immune suppressive response in said vertebrate, connected with (b) a second protein portion analogous to all or a part of an antigen of BHV-1; causing the second protein portion (b) that the vertebrate immune system recognizes the first protein portion (a) and produces an immune response capable of inhibiting the activity of the peptide within the vertebrate when the vertebrate is vaccinated with an effective amount of the protein of fusion.

4. A fusion protein according to claim 3, comprising a portion (a) analogous to all or a part of a GnRH peptide.

5. A fusion protein according to claim 3 wherein the portion (b) is analogous to all or a part of the gD of BHV-1.

6. A polynucleotide molecule comprising a nucleotide sequence encoding a fusion protein according to claim 1 or 3.

7. A vector comprising a polynucleotide molecule according to claim 6.

8. A vector according to claim 7, suitable for the in vitro expression of the fusion protein.

9. A vector according to claim 7, suitable for the in vivo expression of the fusion protein.

10. A transformed cell comprising a polynucleotide molecule comprising in turn a nucleotide sequence encoding a fusion protein according to claim 1 or 4.

11. A double-function vaccine comprising a fusion protein according to claim 1, a vector according to claim 7, or a transformed cell according to claim 10 in an amount effective to inhibit the activity of the peptide from which the portion (a) of the protein is derived. of fusion and to protect with respect to an infection by the pathogen from which the portion (b) of the fusion protein is derived in a vertebrate that endogenously synthesizes the peptide and that can be pathogenically infected by the pathogen, together with a acceptable support for pharmaceutical or veterinary use.

12. A double-function vaccine for inhibiting GnRH activity in cattle animals and for protecting animals of cattle against an infection with BHV-1, comprising a fusion protein according to claim 2, a vector according to claim 7, or a transformed cell according to claim 10 in an amount effective to inhibit the activity of GnRH and protect the animals of cattle against a BHV-1 infection, together with a acceptable support for pharmaceutical or veterinary use.

13. A vaccine for inhibiting the activity of a peptide in a vertebrate comprising a fusion protein according to claim 3, a vector according to claim 7, or a transformed cell according to claim 10 in an amount effective to inhibit the activity of the peptide, together with an acceptable support for pharmaceutical or veterinary use.

14. The use of a fusion protein according to claim 1, of a vector according to claim 7 or of a transformed cell according to claim 10, in the manufacture of a vaccine to inhibit the activity of a peptide synthesized endogenously in a vertebrate and to protect the vertebrate with respect to a pathogenic infection.

15. The use of a fusion protein according to claim 2, of a vector according to claim 7 or of a transformed cell according to claim 10, in the manufacture of a vaccine to inhibit sexual characteristics in a cow and to protect the cow against a BHV-1 infection.

16. The use of a fusion protein according to claim 3, of a vector according to claim 7 or of a transformed cell according to claim 10, in the manufacture of a vaccine to inhibit the activity of a peptide in a vertebrate.

17. The use of a fusion protein according to claim 3, of a vector according to claim 7 or of a transformed cell according to claim 10, in the manufacture of a vaccine to inhibit sexual characteristics, wherein the fusion protein, vector or cell transformed in said vaccine comprises an analogous amino acid sequence, or which encodes an analogous amino acid sequence, to all or a portion of a GnRH peptide. SUMMARY SHEET OF THE INVENTION The present invention provides a fusion protein for producing a double immune response in a vertebrate, which fusion protein comprises: (a) a first protein portion analogous to all or a part of a peptide endogenously synthesized within the vertebrate, the activity whose peptide is to be inhibited within the vertebrate, and whose protein portion is itself incapable of producing an effective immune inhibitory response in said vertebrate; connected with (b) a second protein portion analogous to all or a part of an immunogen from a pathogen, the pathogen of which is capable of infecting the vertebrate pathogenically; causing portion (b) that the vertebrate immune system recognizes the portion (a) and produces a response that: (i) inhibits the activity of the peptide synthesized endogenously within the vertebrate; and (ii) protects the vertebrate with respect to infection by the pathogen, when the vertebrate is vaccinated with an effective amount of the fusion protein; The present invention also provides fusion proteins comprising a protein portion (b) that is a support that is analogous to all or a portion of an antigen of BHV-1, which fusion proteins induce in a vertebrate vaccinated with an effective amount of said fusion protein, an immune response that inhibits activity previously. P00 / 107