JPS62503169A - Molecules containing at least one peptide sequence having an epitope unique to proteins produced by cells infected with malaria parasites, and compositions containing the same - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention] A small number of proteins with unique epitopes produced by cells infected with malaria parasites. Molecules containing one peptide sequence and compositions containing the same The present invention aims to identify epitopes unique to proteins produced by cells infected with malaria parasites. It relates to molecules containing one or more peptide sequences. The invention further includes the molecule a composition for the prevention of malaria, more particularly in which the molecule may constitute one of the active ingredients. Concerning vaccine components.

In particular, Plasmodium ralciparum or Knowlesi - Plasmodium knowlesi or three days A strain of bacteria known as Plasmodium viv■ from sea urchin, a purified strain of the parasite that has been found capable of infecting human hosts. , or from infected cells, especially infected red blood cells, at each stage of disease progression, against malaria. Much research has been done to isolate polypeptide fractions that may have vaccine properties. It is well known that this is being done. Recently, malaria parasite sporozoites derived from the genome and blood form of Plasmodium falciparum (its merozoites) A cloned cDNA bank was established by Ellis J. These bars There were 12 (Palmodium malariae) and 11 (Palmodium falciparum), respectively. including clones capable of expressing polypeptides containing repeating units of amino acids of It is recognized as a thing.

More specifically, the present invention relates to proteins in cells, especially red blood cells, that have been infected by the parasites. Characterized by the presence in the structure of one or several peptide sequences unique to white matter the protein is involved in at least some stages of red blood cell development; It is located in

Molecules of the invention are essentially of the following formula: %formula% (In the formula, X is Glu or Va1%Y is Val, Leu or 1leuSZ is Val or Summer 1e.

A is Glu or Asp, Glu is glutamic acid, Val is valine, lie is isoleucine, Pro is proline, Leu is leucine, and Asp is asparagus. It has at least one nonapeptide sequence represented by (gic acid) It is characterized by

Suitable peptide sequences of the present invention include the following sequences. In addition, the following array In each of 2 to 23, 3 aminoacyl residues in the above first sequence in the vertical direction The groups indicated by are the same as these aminoacyls.

1. GILI GILI Val Mal Glu Glu Val Val Pr.

2, , , , , , , Leu , , , , , , , , , , , , , , , ,.

3. Val , , , Leu , , , , , , , , , , , , , , , , , , , , , , , , , , , ,.

4. Val　, ,Leu　, , , , , , , , , , , , , , .

5,,,,,,,Leu,,,,,,,,,,,,,lie,,,.

6゜,,,,,,Ile,,,,,,,,,,lie,,,’7,,, ,,,,,,,,,,,,,,,,,,,,,,,,,.

8,,,,,,lie,,,,,,,,,,,,,Ile,,.

9, , , , , Leu Ile , , , , , , , , , , , .

10, Mal　, ,Leu　Leu　, ,,,,,,,,lie　,,.

11, Mal, , , , , Leu , , Asp , , Ile 3. .

12, , , , , , , , , , , , , , , , , Ile , .

13,,,,,,,lie　Ile　,,,,,,,,,lie　,,.

14, , , , , Ile Ile , , , , Ala Ile , .

15, Val　,,,lle　,,,,,,,,,,,,lie　,,.

161. ,,Asplie,,,,,,,,,,,,,Ile,,.

17,,,,,,lie　,,,,,,Asp　,,,lie　,,.

1g,,,,,,,,,,,,,,,,,,,,,lie　,,.

19,,,,,. 0. , , , , , , , , , , , , , 8. .

20, , , , , , Leu , , , , , , , , , , Ile , .

21, , 0. ,Leu,,,,,,,,,,,lie,,.

22, , , , , , , , Pro , , , , , , , , , , , , .

23, Glu GIL, Lea, , , , , , , , , , , , , , .

The present invention relates in particular to up to 12 amino acids and to the above formulas, respectively, and to more specific amino acids. from the nonapeptide itself or from 12 amino acids, preferably 9 amino acids. It relates to an oligomer containing a plurality of any one of the above sequences.

It will be appreciated that certain amino acid residues included in the molecular structure of the present invention may have Modification of free reactive groups, especially free carboxyl groups borne on Glu groups, does not inhibit the antigen. Such modifications are possible because they do not change the immunogenicity or, in some cases, the immunity of the entire molecule. It is.

Molecules so modified are of course covered by the claims of the present invention. falls within the scope of protection provided. The carboxyl group may be acylated or esterified.

The peptides of the invention may be produced by conventional methods in the field of peptide synthesis. this The synthesis can be carried out in homogeneous solution or in the solid phase.

For example, llOUBENliEYL's book ``Organic Chemistry Methods'' Method der Organischen Chemie) Yunshi, (E.

Wunsch) vol. 15 I and ■Chaim, Stuttgart 1974 In-solution synthesis methods can be used.

This synthetic method consists of successive condensation of two consecutive aminoacyls in the required order, or or aminoacyl, preformed and already several aminoacyl residues Fragments containing in a specific order, or several pre-generated fragments like this. These aminoacyl or fragments are condensed together. All reactive groups carried on the peptide were prepared according to methods well known in the art of peptide synthesis, especially must intervene in forming the peptide bond after activation of the carboxyl group. One amine group and the other carboxyl group or one carboxyl group and the other All these reactive groups, except the amine group, must be carefully protected beforehand. Variant For example, l-ethyl-3-(3-dimethyl-aminopropyl)-carbodi Using traditional coupling reagents of the carbodiimide type, such as imides, It is also possible to use a coupling reaction that uses The aminoacyl used is When carrying acid groups (particularly in the case of glutamic acid), these reactive groups are e.g. - protected by a butyl ester.

In the case of chained amino acid synthesis, the synthesis begins with the C-terminal amino acid and the adjacent amino acid in the desired sequence. It starts by condensing the noacyl with the corresponding amino acid, and then the N-terminal amino acid is condensed. This can be continued sequentially up to no acids.

According to another preferred method of the invention, R.D. Merryfield (R.D. MERRIFIELD) paper “Solid phase peptide synthesis (Solid phase peptide synthesis)” ptide 5 synthesis)j (J, Am, Chem, Soc, 45.2149-2154) is used.

To generate peptide chains according to Merryfield's method, highly porous A polymer resin is used to fix the first amino acid at the C-terminus of the chain to the resin. this The amino acid is fixed to the resin via the carboxyl group, and the amine group is, for example, t-butyl. protected by a carbonyl group.

Once the first C-terminal amino acid is thus fixed on the resin, the resin can be washed with acid. The protective group for the amine group is removed.

When the protecting group for the amine group is a t-butyloxycarbonyl group, the protecting group It can be removed by treatment of the resin with fluoroacetic acid.

Next, supply the second aminoacyl from the C-terminal aminoacyl residue of the desired sequence. deprotected amino acid of the first C-terminal amino acid fixed to the chain. It binds to the ion group. Preferably, the carboxyl group of the second amino acid is Activated by e.g. dicyclohexylcarbodiimide, the amine group is e.g. - protected by butyloxycarbonyl.

Thus, a site containing two amino acids and having a protected terminal amine group The first portion of the desired peptide chain is obtained. Deprotect the amine group as described above. Then add the third amino acid under the same conditions as the second C-terminal amino acid. can be fixed.

Thus each time a portion of the peptide chain already formed and bound to the resin is Amino acids constituting the peptide chain are sequentially immobilized on the deprotected amine group.

Once the entire desired peptide chain has been formed, each amino acid that makes up the peptide chain The protecting groups are removed from the resin and the peptide is released from the resin, for example with hydrofluoric acid.

The invention further relates to water-soluble oligomers of the monomeric peptides described above. For oligomerization Therefore, the immunity of the monomeric peptides of the invention may be increased. These oligomers are In some cases, it may contain 2 to 25 monomer units (this number shall not be limiting). I can see it.

To perform oligomerization, a total polymerization method commonly used in the peptide field is used. The polymerization can be carried out using as many monomer molecules as necessary to achieve the desired immunity. This is continued until an oligomer or polymer containing the chief is obtained.

A preferred method of oligomerization or polymerization of the monomers is to In addition, it is reacted with a network-forming agent (crosslinking agent).

Other oligomerization or coupling methods can be used as well, e.g. unit through the terminal carboxyl and amine groups in the presence of a heterobifunctional linking agent. A method of sequentially linking molecules can be used.

Similarly, molecules containing one or several nonapeptide motifs as defined above by means of a given nucleic acid containing the appropriate corresponding nucleotide sequence. Genetic engineering techniques using transformed microorganisms can be utilized. correspondence The following nucleotide sequence characterized by a triplet sequence is columns are listed. These sequences are particularly suitable for peptides a) to g) represented by the above formulas. respond. In addition, the nucleotides substituted at points in the sequences 2 to 23 below are the first Vertically identical to the nucleotides of the sequence.

1. GAA GAA GTT GTT GAA GAA GTT GTG CC T2, , , , , , T, A , , , , , , , , , , , , ^2 ．． A3, , T, , , C, A , , A , , , , , , , , , , A 　,,.

4,,T,,,,C,A,,A,,,,,,,,,,,,A, , A5, , , G , , , C, A , , , , , , , G , , , A, A 　,,.

6, , , , , , , A , , , G , , , , , , , , , A, A , ,.

7, , , , , , , C, . , , , , , , , A , , A , .

8, , , G , , , A, A , , , , , , , , , , G A, A , ,.

9, , , , , , , T, A , , , , , , , , , , , , , , , , , , , , , , , , , , , , ,A10,,T,,,,C,A,T,A,,,,,,,,,,A, A,,A11,,T,,,,,,AT,A,,,,,,T,,,, A, A,,.

12, , , , , , , , , , , , , , , , , , , A, A ,,A13,,,,,,,,A,,A,,,,,,,,,,,,,A ,A,,.

14, , , , , , A, , A, , , , , , G, CG A, A ,,A15,,T,,,,A,,,,,,,,,,,G,,,A , A , , A16 , , , , , T A , , , , , , , , , , , , ,A,A,,A17,,,,,,,,A,,,,,,,,,,,, T　、、、　A、A　、、A18゜、、、、、、、、、、、、、、、、 ,,,,,,A,A,,.

19, , , , , , , , , , , , , , , , , , , , , , A 9. A20,,,,,,,T,A,,A,,,,,,,,,A,^　1. A 21, , , , , C, A , , , , , , G , , G A, A , .

22, , , , , , , , CC, , , , G , , , , A , , A23, G AA GAA CTC, , , , , , , , , , , , .

For this purpose, the invention also provides repetitive sequences each containing nine triplets of the type mentioned above. It pertains to one or more nucleic acids. More specifically, up to 1.000 base pairs and each A nucleic acid fragment (fragment) containing 1 to 25 repetitive sequences containing 1 to 25 triplet ).

The present invention further provides for physiologically acceptable non-toxic (natural or synthetic) A bond obtained by covalently bonding the peptide (or the oligomer) of the present invention to a carrier molecule. Concerning the merger. The reactive groups are complementary to each other supported on the carrier molecule and the peptide, respectively. It is a reactive group. Examples of suitable reactive groups are given below.

Examples of carrier molecules or polymeric carriers involved in the composition of the conjugate of the present invention include natural protein Paku, e.g., tetanus anatoxin, ovalbumin, serum albumin, hemosia There are Min et al.

As a synthetic polymer carrier, for example, polylysine or poly(D-L-alanine)-poly (L-lysine).

Other polymeric carriers that can be used are also described in the literature, and the molecular weight of these polymeric carriers is generally greater than 20,000.

The conjugates of the invention can be synthesized using known methods, such as Infec. t, and Immunity) J, 33.193-198 (1981). FRANTZ and ROBERTSON Methods, or “Applied and Environm micro-bjology) J, October 1981, vol. 42, 4 No. 1, pp. 611-614 using the peptides and suitable carrier molecules. The method of E. Kaufman (P, E. KAUFFMAN) may be used.

In practice, it is advantageous to use as binders the following compounds, which are given by way of non-limiting example: be. Glutaraldehyde, ethyl chloroformate, water-soluble carbodiimide For example, HCL-N-ethyl-N.

(3-dimethylamino-propyl-carbodiimide, diisocyanate, bis- Diazobenzidine, di- and trichloro-5-)riazine, cyanogen bromide, Nzoquinone. Additionally, the Scandinavian Journal of Immunology (Sc and, J. [mmunol,), 1978.8, 7-23 Hage (AVR AMEAS, TERNYNCK%GUESDON) J: The binder described in g is also advantageous. be.

Reacting one or more reactive functional groups of the peptide with one or more reactive functional groups of the carrier molecule. Any suitable attachment method may be used. Types of bonds used in protein synthesis agents, such as 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide , N-hydroxy-benzotriazole, etc., which can undergo a bonding reaction. Preference is given to using xyl and amine functions. Also, especially with peptides When bonding the amine groups supported on each carrier molecule to each other, glutamate Use Rudehyde.

Additional features of the invention will become apparent from the following description based on the accompanying drawings.

In the following description, a polypeptide containing multiple nonapeptide sequences having the above structure is obtained. Explain the conditions under which

Figure 1 shows the modified plastic used to produce the polypeptide containing the nonapeptide sequence. FIG. 2 is a schematic illustration of the structure of Smid, and FIG. 2 shows a nucleic acid capable of producing the polypeptide. It shows the partial structure of ``Enzymology (Methods in E Maxam and Gilbert described in A65.499-560 (Maxam et G11bert) (1980) was used.

Plasmodicm falciparum National Microbial Culture Center (C,N,C,N) on December 2, 1982. Genomic DNA previously isolated from the bacterial strain (deposited on the 3rd with accession number FUPCI-212) Deoxyribonuclease (DNAse) on the order of 200 to 1,000 base pairs Cut into pieces. These fragments were introduced into the vector pUK270 with the structure shown in Figure 1. I entered. Plasmid I) UK270 was derived from Koenen (M, KOENEW) et al. (19 82) Plasmid pUK23 described by EMBO111509-512 0 sequence. Plasmid pUK230 and plasmid Do pI! The difference between 270 and 270 is that the 5° of the gene encoding β-galactosidase There is a polylinker sequence inserted in the section. This synthetic nucleotide sequence (polylinker) introduces a phase shift within two genes. The effect is genetic The purpose is to change Z, Y, and A to the phenotype Z-, Y-1A-.

Before introducing the genomic DNA fragment of Plasmodium falciparum into the vector, The Pst-1 site was cleaved to form an oligo-G single strand at the end. DNA The strand itself is pre-equipped with an oligo-C terminus so that the DNA strand is can be combined with a ter. When a DNA fragment is introduced into the Pst-I site under the above conditions, cloning occurs. can be obtained. This clone has the same gene 2 as M Koenen (M, KOE It is aligned with the promoter under the conditions described by NIJ), making it lactose positive. Become. These clones are 5-bromo-4-chloro-3-indolyl-β-D- Galactosidase (X-gal) or isopropyl-1-thio β-D-galac detected as blue colored in indicator plaques containing toside (IPTG), or , detected as lactose-positive colonies on lactose agar. In contrast, it does not contain fragments. The vector is colored pale blue with the same indicator plaque and is characterized by a lactose-negative phenotype. Ru. Therefore, the lactose-positive recombinant is a chimeric protein containing the main part of β-galactosidase. Express the inserted DNA as the N-terminal part of the park.

Antigen-producing clones were identified using the immunoenzymatic method of Kohnen et al. Lactose positive 10,000 clones were isolated and screened using various antisera under the conditions described below. -ning.

First, falciparum malaria was denatured with a detergent known as Triton x 100. Antibodies from rabbits immunized with protozoan partial conjugates (merozoites) were used. 2 times contact Serum was collected from rabbits after inoculation. The IgG fraction was isolated and used for immunoenzyme detection. .

In the second screening, we selected locally-born individuals with high antibody titers against Plasmodium falciparum. A mixture of antisera collected from four individuals was used. Escherichia coli extract ground before use The anti-E. coli antibodies were completely removed from the antiserum by contacting it with the serum and diluted 25 times. .

In detail, the test was performed using the following procedure.

Lactose-positive clones were cultured on lactose-containing agar in glass veterinary dishes and Bacteriolysis was performed under the conditions described above. Some cultures are grown on nitrocellulose on lactose-containing agar. Cultured on a lurose filter. Clothe any growing colonies, especially the filter, at room temperature. Bacteria were lysed by contacting with loform for 1 minute. Expose the filter to air for 5 minutes at 37°C. to remove chloroform. Next, the filter was coated with polyvinylchloride that had been previously coated with antibodies. was brought into contact with nil sheet. Proceedings of the National Antibody Coating Academic Science (Proc, Natl, Acad, Sci,), USA , 75.2746-2749 (197g) A method such as OME) is used. Dissolve this in a cleaning solution as described by Kohnen et al. saturated with decomposed serum albumin. This cleaning solution is further added to 0.5% (vol/vol) Contains 100 Tritons. After immune absorption, take out the sheet and place Triton x1 00 for 20 minutes. Remove any cell debris that is still attached to the syringe. was removed with a jet of the same solution used. Next, place the sheet in new solution and any debris will be visible. Keep f2 until it runs out.

Next, immune complexes were identified using the method of M. Coenen et al.

In this way, 36 positive clones that reacted with rabbit antibodies were isolated. correspondence colonies were screened under similar conditions. However, this time, the above-mentioned The prepared human serum was used.

One of these clones reacted particularly strongly with anti-human serum.

The products of this clone were subjected to sodium dodecyl sulfate acrylamide gel electrophoresis. 116 kilodalton peptide corresponding to β-galactosidase. It was discovered that the 150 kilodalton band does not exist and a new band exists at 150 kilodaltons. Ta. This band is isolated in relatively large amounts in the cytoplasmic fraction of E. coli and is rare in the membrane. .

This result is consistent with the size of the approximately 620 base pair insert incorporated into the vector. I did it. This insert was sequenced in two directions using the method of Maxam and Gilbert. Established. As a result, especially regarding the structure of the 620 base pair terminal sequence, Full-length insert derived from the Plasmodium genome [9 amino acid repeat polypeptide] Glu Glu Leu Val GILI GILI Mal Vat Pr It was estimated that it contains a 27 base pair repeat sequence encoding o. Of course these sequences There are some changes within. Especially isoleucine, leucine and valine amino acids. No acids can be substituted for each other. As a result of observations regarding these sequences, proline This suggests the existence of an alpha helix containing a blockade. Characteristic DNA sequence and its encoded The peptide sequence obtained is clear from FIG.

The polypeptide thus expressed exhibits the characteristics of the polypeptide of the invention.

The recognition of the chimeric protein by human antibodies is due to the presence of characteristic antigenic determinants. This supports the hypothesis that it is a human immunogen. E Southern (E, 5 OUT HERN) “Southern plot method J (1975) J, Mo1. Biol, 98 503, sequence analysis of genomic DNA revealed that the relevant fragment was caused by the old ndIII enzyme. A band of approximately 20-25 kb was detected after digestion. using poly-A″ RNA. -than plot method (P, F, Tl (OMAS (1980) Proc, Na tl, Acad, Sci, 77.5201) but 7 to 9 kb of Metsenger R NA was successfully identified.

Polypeptides comprising repetitive peptide sequences of the invention have immunological properties. Especially β-ga Lactosidase-fragment chimeric molecule (β-galactosidase acts as a carrier) ) antibodies formed against terminal red blood cells infected with Plasmodium falciparum. It was found that it can react with certain antigens. This reaction produces a characteristic image using indirect immunofluorescence. It is detected by The same immunofluorescence image shows multiple P. falciparum strains. (PaloALto, NP54, ItG2G1). to this All immunofluorescence methods known to those skilled in the art, e.g. ELISA, immunofluorescence It is believed that polypeptide sequences can be used in the diagnosis of Plasmodium falciparum by et al. good.

Plasmodium falciparum antigen recognized by serum a under the above experimental conditions (1% Plasmodium falciparum was extracted using a 6.5% 5DS-polyacrylamide gel. (subjected to pneumophoresis) has a molecular weight of approximately 250,000 daltons.

Due to its antigenicity, the molecules of the invention are therefore particularly useful as a result of the infectious action of malaria parasites. Acts particularly on proteins produced during red blood cell development, and more specifically during infection. Used as a reagent to detect antibodies that act against proteins associated with blood cell membranes. Can be used. Some of these proteins include an editor sometimes found in cerebral forms of malaria. Some may involve the fixation of infected red blood cells to capillary endothelial cells. This phenomenon affects patients can cause malaria thrombosis, which is often fatal.

Conversely, the peptides or oligopeptides of the present invention can be used by immunizing animals such as rabbits. It can also be used to produce antibodies in vivo. These antibodies At some stage during the progression of a protozoan infection, membrane proteins of red blood cells or exposed on the surface of the membrane can be used to diagnose in vitro the appearance of proteins that

The present invention also provides immune agents (v) against these particularly malignant immunopathological phenomena. This will pave the way for the development of new ingredients for In particular, the above results , with or without covalently bonding the peptide or oligopeptide of the invention to a support. It can be used as a component to prevent the fixation of infected red blood cells to the walls of knitted capillaries. and make it possible. Therefore, the molecule of the present invention is an active ingredient of a vaccine that acts against malaria. may constitute one of the following.

The present invention therefore provides a peptide of the present invention, an oligomer of this peptide, or a peptide of this invention. The peptide or oligomer bound to the carrier molecule (conjugate) is Included with the vehicle and if necessary, another effective substance that has an immune effect against malaria. It also relates to compositions formulated in vaccine form that also contain the following ingredients:

An advantageous pharmaceutical composition is an injectable composition containing an effective amount of at least one product of the invention. Consists of solution, suspension or liposome. Preferably, these solutions, suspensions or liquids Posomes are formed in a sterile isotonic aqueous phase, preferably saline or glucose-containing aqueous phase. do.

The invention is more particularly suitable for administration by intradermal injection, intramuscular injection, subcutaneous injection or scarification. Suitable formulations include suspensions, solutions or liposomal forms as described above.

The invention further provides for alternative methods of administration, especially oral or rectal administration, or especially ocular, nasal, pulmonary administration. or a pharmaceutical composition suitable for administration in the form of an aerosol in contact with mucous membranes, such as the mucous membranes of the vagina. It also applies to products.

The invention therefore provides for the administration of at least one product of the invention orally, intraocularly or nasally. Pharmaceutically acceptable solid or liquid excipients suitable for the formation of intrarectal dosage forms; excipients suitable for forming dosage forms or gelatinous excipients suitable for intravaginal administration; It relates to pharmaceutical compositions comprising the above in combination with any of the above. The invention also particularly suitable for administration to the ocular or nasal mucosa, and containing at least one conjugate of the invention, etc. It also relates to liquid compositions.

The vaccine composition of the present invention further comprises polyvinyl- It is advantageous to include a vehicle such as pyrrolidone. Polyvinyl-pyrrolidone replacement has the conventional meaning, i.e. facilitates the absorption of a drug or the action of a drug in vivo. Any other adjuvant in the sense of a substance that makes the action more active may be used. stomach. Other adjuvants of the latter type include, for example, carboxymethyl-cellulose. base, aluminum hydroxide, aluminum phosphate, or as known to those skilled in the art. All other adjuvants in the formulation are included. The compositions of the invention can be used for immunization if necessary. Also included are chemical adjuvants, particularly those of the muramyl peptide type.

The present invention is of course not limited to the non-limiting embodiments described above, and does not depart from the scope of the following claims. Various modifications may be made by those skilled in the art. In particular, intervening peptide sequences of the invention Some amino acids also have equivalent functions. may be substituted with isot6riques amino acids. -For example: One or more substitutions may be made.

- replacing Glu with Asp or Gin, - Replace Val with Ala or Gly, -Replace Leu with Ala, etc.

Not only the peptides obtained by such substitutions but also the peptides themselves or these peptides Even if oligomers or conjugates formed using more particularly consisting of equivalents of the claimed peptides. It turns out.

More specifically, the present invention relates to "chimeric proteins" which can be obtained by genetic engineering technology. It also concerns “white matter.” These chimeric proteins contain peptide fragments other than β-galactosidase. one or more peptide sequences incorporated into or attached to a fragment, especially a nonape of the invention. may include putide. This peptide fragment is preferably an immunizing agent of the peptide sequence of the invention. It has a molecular weight sufficient to enhance genicity, and can inhibit the expression of desired immunity from an immunological point of view. We don't get along.

Nucleotide sequences corresponding to the above peptide sequences can be determined using known molecular hybridizers. According to the tion technique, B. fufurciparum (P.

falciparum) can be used as a substance (reagent) for diagnosis.

Similarly, the peptides encoded by these nucleotide sequences are The patient's serum contains antibodies that act against the aforementioned proteins associated with the membranes of infected blood cells. can be used to detect These peptides, on the other hand, are particularly important for antibodies in the animal body. These antibodies can also be used to produce malaria parasites, particularly in immunological in vitro by bringing them into contact with potentially infectious blood cells under conditions that allow a reaction to occur. used for detection in

FIG. 1.

)! 1ndTiXba　]″8all　PsLI　BamHI　EcoRUl, , GAA 688 GTT GTT GAA GAA GTT 13TG CCT ICL, T,,,C-he Ding,he,,,-,,+++ A, A++8.

11,,T,,,,,,8■,Δ,-,,T-,,mu,to 0. .

20, , , -, , T, 8 6. Mu 01160001. 8-8 1. 821 , +++ +++ c, 8, -, , -4G, J 8, A +++---6 tu　Vat　V(11Pr.

S--,,-,-Leu,,,-,,-,-+++lie-,.

60. ,,+++Ite-1,-6,-,,-1,Ile-,.

9, +++, -, Leu Ile +++, -6+++ 0. , -0゜1 0, V(11-,, Leu Leu +++ 0., 0., lie -1°11 , Vat , , -1, Leu -, , 8 sp +++ lie -, -11+ ＋＋ ＋＋＋ ＋＋＋ ＋＋＋ ＋＋＋ ＋＋＋ ＋＋＋ Itoe, -0゜1 3, +++ -,, Ile l1e-,, +++ +++ Ile, -.

5- Vat +++ lie +++,,, -,,+++ Ite++ +6, +++ 8 sp Ile +++ +++ +++ +++ l1e-.

17,,,,-,,lle,,,,,,As,,,,Ile,,- 20, +++ +++ Leu +++ -, -+++,, -11e +++ 2], ++, +++ Leu 0000. ,-,,-0. Ile-,.

21-,-,,,-,-Pro-,,,,,,,-,+++,,.

international search report AR'EXTo-:'EINTEEIAT:ON input LSEARCHREPORT ONUS-A-444203110104/84 None

Claims (1)

[Claims] 1. Epitopes unique to proteins produced by cells infected with malaria parasites A molecule comprising at least one peptide sequence having The following formula: [There is an array] [Wherein, X is Glu or Val, Y is Va1, Leu or lleu, Z is Val or lle and A are Glu or Asp. ] Child. 2. The peptide sequence is as follows: 1. [There is an array] 2. [There is an array] 3. [There is an array] 4. [There is an array] 5. [There is an array] 6. [There is an array] 7. [There is an array] 8. [There is an array] 9. [There is an array] 10. [There is an array] 11. [There is an array] 12. [There is an array] 13. [There is an array] 14. [There is an array] 15. [There is an array] 16. [There is an array] 17. [There is an array] 18. [There is an array] 19. [There is an array] 20. [There is an array] 21. [There is an array] 22. [There is an array] 23. [There is an array] Molecule according to claim 1, characterized in that it is one of: It consists of a monomeric peptide containing 3.9 amino acids, and this peptide has the above sequence. Molecule according to claim 1 or claim 2, characterized in that it has one of the following. 4. Nonapeptide having a structure corresponding to the structure of any one of the above sequences The molecule according to claim 1 or claim 2, characterized in that the molecule consists of 5. It is characterized in that it contains a plurality of repeated sequences corresponding to one or more sequences in the above-mentioned sequences. A molecule according to claim 1 or claim 2 characterized in that it is characterized by: 6. It consists of an oligomer of one or more sequences in the above sequence, and this oligomer is composed of two or more sequences. Claim 1 or Claims characterized in that it contains 25 monomer units from Molecules according to Box 2. 7. According to any one of claims 1 to 6, the device is fixed to a support. molecule of. 8. A molecule according to any of claims 1 to 7 together with an acceptable pharmaceutical vehicle. An immunogenic composition comprising: 9. comprising a molecule according to any of claims 1 to 7 together with other immunogenic components. A vaccine composition acting against malaria. 10. one or more of the peptide sequences set forth in claim 1 or claim 2. Encoding nucleotide sequence. 11. Use of a molecule according to any of claims 1 to 7 as a diagnostic reagent. 12. Use of the nucleotide sequence according to claim 10 as a diagnostic reagent.