WO2001038379A1 - A new polypeptide-human ribosomal protein l23 and the polynucleotide encoding it - Google Patents

Abstract

The present invention discloses a new polypeptide- human ribosomal protein L23, the polynucleotide encoding it and a method producing the polypeptide by recombinant DNA technology. The present invention further discloses a method using the polypeptide to treat various disorders, e.g. malignant neoplasm, hematopathy, HIV infection and immunological diseases and various inflammations etc. The present invention also discloses an agonist of the polypeptide and its therapeutic use. The present invention further discloses the use of the polynucleotide encoding the new human ribosomal protein L23.

Description

 Explanation book

 A new polypeptide-human ribosomal protein L23 and polynucleotides encoding this polypeptide

 The present invention belongs to the field of biotechnology. Specifically, the present invention describes a new polypeptide, human ribosomal protein L23, and a polynucleotide sequence encoding the polypeptide. The invention also relates to a preparation method and application of the polynucleotide and the polypeptide. Background technique

 Ribosomes are organelles that synthesize proteins. The main components are proteins and ribosomal RNA. Ribosome proteins are very conserved and play an important role in the process of ribosomal translation. For example: they play an important role in folding rRNA into a functional three-dimensional structure; in protein synthesis, a series of changes in the spatial conformation of the ribosome, some Ribosomal proteins may "fine-tune" the conformation of the ribosome; at the binding site of the ribosome, it may even play a catalytic role, and the ribosome protein functions together with rRNA. L22 ribosomal protein is one of the five ribosomal proteins necessary for early folding of 23S rRNA. L22 is distributed on the 50S ribosomal subunit, close to the active site of peptidyl transferase. The L22 protein includes a small c + β domain and a 30A hairpin structure protruding from the P subunit. Most of the surface of this protein has the potential to interact with rRNA. Similar structures also exist in some other RNA-binding proteins, suggesting that they may have the same origin. Deletion or inappropriate chemical modification of ribosomal proteins, or mutations in ribosomal protein genes, will affect the function of ribosomes and reduce the activity of peptide synthesis, leading to various serious consequences, such as the inability to maintain cell membrane structures and cells. Metabolic disorders, reduced cell surface antibodies, cell growth halt, and even death, etc., cause various diseases, such as developmental disorders, metabolic diseases, immune diseases, cell death, and so on. The present invention has 33% homology at the protein level with the L22 protein found in aquatic bacteria, so it is believed that the base of the present invention is because it encodes a gene similar to the L22 protein and has similar biological functions. The present invention is named human ribosomal protein L23.

As described above, the human ribosomal protein L23 protein plays an important role in regulating important functions of the body such as cell division and embryonic development, and it is believed that a large number of proteins are involved in these regulatory processes, so there has been a need in the art to identify more involved in these processes. Human ribosomal protein L23 protein, especially the amino acid sequence of this protein is identified. Isolation of the new human ribosomal protein L23 protein encoding gene has also been used to determine the role of this protein in health and disease states. Use provides the foundation. This protein may form the basis for developing diagnostic and / or therapeutic drugs for diseases, so isolating its coding DNA is important. Disclosure of invention

 It is an object of the present invention to provide isolated novel polypeptides-human ribosomal protein L23 and fragments, analogs and derivatives thereof.

 Another object of the invention is to provide a polynucleotide encoding the polypeptide.

 Another object of the present invention is to provide a recombinant vector containing a polynucleotide encoding human ribosomal protein L23.

 Another object of the present invention is to provide a genetically engineered host cell containing a polynucleotide encoding human ribosomal protein L23.

 Another object of the present invention is to provide a method for producing human ribosomal protein L23.

 Another object of the present invention is to provide an antibody against the polypeptide-human ribosomal protein L23 of the present invention. Another object of the present invention is to provide mimic compounds, antagonists, agonists, and inhibitors against the human ribosomal protein L23 of the polypeptide of the present invention.

 Another object of the present invention is to provide a method for diagnosing and treating diseases associated with abnormalities of human ribosomal protein L23.

 The present invention relates to an isolated polypeptide, which is of human origin, and includes: a polypeptide having the amino acid sequence of SEQ ID D. 2 or a conservative variant, biologically active fragment or derivative thereof. Preferably, the polypeptide is a polypeptide having the amino acid sequence of SEQ ID NO: 2.

 The invention also relates to an isolated polynucleotide comprising a nucleotide sequence or a variant thereof selected from the group consisting of:

(a) a polynucleotide encoding a polypeptide having the amino acid sequence of SEQ ID No. 2;

 (b) a polynucleotide complementary to polynucleotide (a);

 (c) A polynucleotide having at least 70% identity to a polynucleotide sequence of (a) or (b).

 More preferably, the sequence of the polynucleotide is one selected from the group consisting of: (a) a sequence having positions 20 to 640 in SEQ ID NO: 1; and (b) a sequence having 1- in SEQ ID NO: 1 707-bit sequence.

 The invention further relates to a vector, in particular an expression vector, containing the polynucleotide of the invention; a host cell genetically engineered with the vector, including a transformed, transduced or transfected host cell; and a method comprising culturing said Host cell and method of preparing the polypeptide of the present invention by recovering the expression product.

The invention also relates to an antibody capable of specifically binding to a polypeptide of the invention. The invention also relates to a method for screening compounds that mimic, activate, antagonize or inhibit the activity of human ribosomal protein L23 protein, which comprises utilizing the polypeptide of the invention. The invention also relates to compounds obtained by this method.

 The invention also relates to a method for detecting a disease or susceptibility to disease associated with abnormal expression of human ribosomal protein L2 3 protein in vitro, which comprises detecting a mutation in the polypeptide or a polynucleotide sequence encoding the same in a biological sample, or detecting a biological The amount or biological activity of a polypeptide of the invention in a sample.

 The present invention also relates to a pharmaceutical composition, which comprises a polypeptide of the present invention or a mimetic thereof, an activator, an antagonist or an inhibitor, and a pharmaceutically acceptable carrier.

 The present invention also relates to the use of the polypeptides and / or polynucleotides of the present invention in the preparation of a medicament for treating cancer, developmental or immune diseases, or other diseases caused by abnormal expression of human ribosomal protein L23.

 Other aspects of the invention will be apparent to those skilled in the art from the disclosure of the techniques herein.

 The following terms used in this specification and claims have the following meanings unless specifically stated otherwise:

 "Nucleic acid sequence" refers to oligonucleotides, nucleotides or polynucleotides and fragments or parts thereof, and can also refer to genomic or synthetic DNA or RNA, which can be single-stranded or double-stranded, representing the sense strand or Antisense strand. Similarly, the term "amino acid sequence" refers to an oligopeptide, peptide, polypeptide or protein sequence and fragments or portions thereof. When the "amino acid sequence" in the present invention relates to the amino acid sequence of a naturally occurring protein molecule, such "polypeptide" or "protein" does not mean to limit the amino acid sequence to a complete natural amino acid related to the protein molecule .

 A protein or polynucleotide "variant" refers to an amino acid sequence having one or more amino acids or nucleotide changes or a polynucleotide sequence encoding it. The changes may include deletions, insertions or substitutions of amino acids or nucleotides in the amino acid sequence or nucleotide sequence. Variants can have "conservative" changes, in which the substituted amino acid has a structural or chemical property similar to that of the original amino acid, such as replacing isoleucine with leucine. Variants can also have non-conservative changes, such as replacing glycine with tryptophan.

 "Deletion" refers to the deletion of one or more amino acids or nucleotides in an amino acid sequence or nucleotide sequence. "Insertion" or "addition" refers to an alteration in the amino acid sequence or nucleotide sequence that results in an increase in one or more amino acids or nucleotides compared to a naturally occurring molecule. "Replacement" refers to the replacement of one or more amino acids or nucleotides with different amino acids or nucleotides.

 "Biological activity" refers to a protein that has the structure, regulation, or biochemical function of a natural molecule. Similarly, the term "immunologically active" refers to the ability of natural, recombinant, or synthetic proteins and fragments thereof to induce a specific immune response and to bind specific antibodies in a suitable animal or cell.

An "agonist" refers to a molecule that, when combined with human ribosomal protein L23, can cause the protein to change, thereby regulating the activity of the protein. Agonists can include proteins, nucleic acids, carbohydrates, or any other binding human Molecule of ribosomal protein L23.

 An "antagonist" or "inhibitor" refers to a molecule that, when combined with human ribosomal protein L23, can block or regulate the biological or immunological activity of human ribosomal protein L23. Antagonists and inhibitors may include proteins, nucleic acids, carbohydrates or any other molecule that can bind to human ribosomal protein L23.

 "Regulation" refers to a change in the function of human ribosomal protein L23, including an increase or decrease in protein activity, a change in binding properties, and any other biological, functional, or immune properties of human ribosomal protein L23.

 "Substantially pure" means substantially free of other proteins, lipids, sugars or other substances with which it is naturally associated. Those skilled in the art can purify human ribosomal protein L23 using standard protein purification techniques. The substantially pure human ribosomal protein L23 produces a single main band on a non-reducing polyacrylamide gel. The purity of human ribosomal protein L23 peptide can be analyzed by amino acid sequence.

 "Complementary" or "complementary" refers to a polynucleotide that naturally binds by base-pairing under conditions of acceptable salt concentration and temperature. For example, the sequence "C-T-G-A" can be combined with the complementary sequence "G-A-C-T". The complementarity between two single-stranded molecules can be partial or complete. The degree of complementarity between nucleic acid strands has a significant effect on the efficiency and strength of hybridization between nucleic acid strands.

 "Homology" refers to the degree of complementarity and can be partially homologous or completely homologous. "Partial homology" refers to a partially complementary sequence that at least partially inhibits hybridization of a fully complementary sequence to a target nucleic acid. This inhibition of hybridization can be detected by performing hybridization (Sou thern blot or Nor thern blot, etc.) under conditions of reduced stringency. Substantially homologous sequences or hybridization probes can compete and inhibit the binding of completely homologous sequences to the target sequence under conditions of reduced stringency. This does not mean that the conditions of reduced stringency allow non-specific binding, because the conditions of reduced stringency require that two sequences bind to each other as a specific or selective interaction.

 "Percent identity" refers to the percentage of sequences that are identical or similar in the comparison of two or more amino acid or nucleic acid sequences. The percent identity can be determined electronically, such as by the MEGAL IGN program (Lasergene sof tware package, DNASTAR, Inc., Madi son Wis.). The MEGALIGN program can compare two or more sequences according to different methods such as Cluster method (Higgins, D. G. and P. M. Sharp (1988) Gene 73: 237-244). The Clus ter method arranges groups of sequences into clusters by checking the distance between all pairs. The clusters are then allocated in pairs or groups. The percent identity between two amino acid sequences such as sequence A and sequence B is calculated by the following formula:

 Number of matching residues between sequence A and sequence B

 X 100 Number of residues in sequence A-number of spacer residues in sequence A-number of spacer residues in sequence B

Phases between nucleic acid sequences can also be determined by the Clus ter method or by methods known in the art such as Jo tun He in. Percent homosexuality (He in J., (1990) Methods in emzumol ogy 183: 625-645).

 "Similarity" refers to the degree to which the amino acid residues at the corresponding positions are replaced by the same or conservative substitutions in the alignment between amino acid sequences. Amino acids used for conservative substitutions, for example, negatively charged amino acids may include aspartic acid and glutamic acid; positively charged amino acids may include lysine and arginine; having an uncharged head group is Similar hydrophilic amino acids may include leucine, isoleucine and valine; glycine and alanine; asparagine and glutamine; serine and threonine; phenylalanine and tyrosine.

 "Antisense" refers to a nucleotide sequence that is complementary to a particular DNA or RNA sequence. "Antisense strand" refers to a nucleic acid strand that is complementary to the "sense strand".

 "Derivative" refers to a chemical modification of HFP or a nucleic acid encoding it. This chemical modification may be the replacement of a hydrogen atom with an alkyl, acyl or amino group. Nucleic acid derivatives can encode polypeptides that retain the main biological properties of natural molecules.

"Antibody" refers to a complete antibody molecule and its fragments, such as Fa, (& 1) ') ₂ and 1 ^, which can specifically bind to the epitope of human ribosomal protein L2 3.

 A "humanized antibody" refers to an antibody in which the amino acid sequence of a non-antigen binding region is replaced to become more similar to a human antibody, but still retains the original binding activity.

 The term "isolated" refers to the removal of a substance from its original environment (for example, its natural environment if it occurs naturally). For example, a naturally occurring polynucleotide or polypeptide is not isolated when it is present in a living thing, but the same polynucleotide or polypeptide is separated from some or all of the substances that coexist with it in the natural system. Such a polynucleotide may be part of a vector, or such a polynucleotide or polypeptide may be part of a composition. Since the carrier or composition is not a component of its natural environment, they are still isolated. As used herein, "isolated" refers to the separation of a substance from its original environment (if it is a natural substance, the original environment is the natural environment). For example, polynucleotides and polypeptides in a natural state in a living cell are not isolated and purified, but the same polynucleotides or polypeptides are separated and purified if they are separated from other substances existing in the natural state. .

 As used herein, "isolated human ribosomal protein L23" means that human ribosomal protein L2 3 is substantially free of other proteins, lipids, sugars, or other substances with which it is naturally associated. Those skilled in the art can purify human ribosomal protein L23 using standard protein purification techniques. Substantially pure polypeptides can produce a single main band on a non-reducing polyacrylamide gel. The purity of the human ribosomal protein L23 polypeptide can be analyzed by amino acid sequence.

The present invention provides a new polypeptide, human ribosomal protein L23, which is basically composed of the amino acid sequence shown in SEQ ID NO: 2. The polypeptide of the present invention may be a recombinant polypeptide, a natural polypeptide, a synthetic polypeptide, A recombinant polypeptide is preferred. The polypeptides of the present invention can be naturally purified products or chemically synthesized products, or can be produced from prokaryotic or eukaryotic hosts (eg, bacteria, yeast, higher plants, insects, and mammalian cells) using recombinant techniques. Depending on the host used in the recombinant production protocol, the polypeptide of the invention may be glycosylated, or it may be non-glycosylated. Polypeptides of the invention may also include or exclude starting methionine residues.

 The invention also includes fragments, derivatives and analogs of human ribosomal protein L23. As used in the present invention, the terms "fragment", "derivative" and "analog" refer to a polypeptide that substantially maintains the same biological function or activity of the human ribosomal protein L23 of the present invention. A fragment, derivative or analog of the polypeptide of the present invention may be: (I) a kind in which one or more amino acid residues are substituted with conservative or non-conservative amino acid residues (preferably conservative amino acid residues), and the substitution The amino acid may or may not be encoded by a genetic codon; or (Π) such a type in which one or more amino acid residues are substituted with other groups to include a substituent; or (III) such One, in which the mature polypeptide is fused to another compound (such as a compound that prolongs the half-life of the polypeptide, such as polyethylene glycol); or (IV) such a polypeptide sequence in which the additional amino acid sequence is fused into the mature polypeptide ( Such as the leader sequence or secreted sequence or the sequence used to purify this polypeptide or protease sequence) As explained herein, such fragments, derivatives and analogs are considered to be within the knowledge of those skilled in the art.

 The present invention provides an isolated nucleic acid (polynucleotide), which basically consists of a polynucleotide encoding a polypeptide having the amino acid sequence of SEQ ID NO: 2. The polynucleotide sequence of the present invention includes a nucleotide sequence of SEQ ID NO: 1. The polynucleotide of the present invention is found from a cDNA library of human fetal brain tissue. It contains a polynucleotide sequence of 707 bases in length and its open reading frame (20-640) encodes 206 amino acids. Based on the amino acid sequence homology comparison, it was found that this polypeptide has 33% homology with the L22 protein, and it can be deduced that the human ribosomal protein L23 has a similar structure and function to the L22 protein.

 The polynucleotide of the present invention may be in the form of DNA or RNA. DNA forms include cDM, genomic DNA or synthetic DNA. DNA can be single-stranded or double-stranded. DNA can be coding or non-coding. The coding region sequence encoding the mature polypeptide may be the same as the coding region sequence shown in SEQ ID NO: 1 or it may be a degenerate variant. As used herein, a "degenerate variant" refers to a nucleic acid sequence encoding a protein or polypeptide having SEQ ID NO: 2 but different from the coding region sequence shown in SEQ ID NO: 1 in the present invention.

The polynucleotide encoding the mature polypeptide of SEQ ID NO: 2 includes: only the coding sequence of the mature polypeptide; the coding sequence of the mature polypeptide and various additional coding sequences; the coding sequence of the mature polypeptide (and optional additional coding sequences) Coded sequences) and non-coded sequences.

 The term "polynucleotide encoding a polypeptide" refers to a polynucleotide that includes the polypeptide and a polynucleotide that includes additional coding and / or non-coding sequences.

 The invention also relates to variants of the polynucleotides described above, which encode polypeptides or fragments, analogs and derivatives of polypeptides having the same amino acid sequence as the invention. This polynucleotide variant can be a naturally occurring allelic variant or a non-naturally occurring variant. These nucleotide variants include substitution variants, deletion variants, and insertion variants. As known in the art, an allelic variant is an alternative form of a polynucleotide that may be a substitution, deletion, or insertion of one or more nucleotides, but does not substantially change the function of the polypeptide it encodes .

 The invention also relates to a polynucleotide that hybridizes to the sequence described above (having at least 50%, preferably 70% identity between the two sequences). The present invention particularly relates to polynucleotides that can hybridize to the polynucleotides of the present invention under stringent conditions. In the present invention, "strict conditions" means: (1) hybridization and elution at lower ionic strength and higher temperature, such as 0.2xSSC, 0.1 ° /. SDS, 60 ° C; or (2) adding denaturants during hybridization, such as 50% (v / v) formamide, 0.1% calf serum / 0.1% F ico ll, 42 ° C, etc .; or (3) Hybridization occurs only when the identity between the two sequences is at least 95%, and more preferably 97%. Moreover, the polypeptide encoded by the hybridizable polynucleotide has the same biological function and activity as the mature polypeptide shown in SEQ ID NO: 2.

 The invention also relates to nucleic acid fragments that hybridize to the sequences described above. As used in the present invention, a "nucleic acid fragment" contains at least 10 nucleotides in length, preferably at least 20-30 nucleotides, more preferably at least 50-60 nucleotides, and most preferably at least 100 nuclei. Glycylic acid or more. Nucleic acid fragments can also be used in nucleic acid amplification techniques, such as PCR, to identify and / or isolate polynucleotides encoding human ribosomal protein L23.

 The polypeptides and polynucleotides in the present invention are preferably provided in an isolated form and are more preferably purified to homogeneity.

 The specific polynucleotide sequence encoding the human ribosomal protein L23 of the present invention can be obtained by various methods. For example, polynucleotides are isolated using hybridization techniques well known in the art. These techniques include, but are not limited to: 1) hybridization of probes to genomic or cDNA libraries to detect homologous polynucleotide sequences, and 2) antibody screening of expression libraries to detect cloned polynucleosides with common structural characteristics Acid fragments.

 The DM fragment sequence of the present invention can also be obtained by the following methods: 1) isolating the double-stranded DNA sequence from the genomic DNA; 2) chemically synthesizing the DNA sequence to obtain the double-stranded DNA of the polypeptide.

Of the methods mentioned above, genomic DNA isolation is the least commonly used. Direct chemical synthesis of DNA sequences is often the method of choice. The more commonly used method is the isolation of cDNA sequences. Isolation of cDNA of interest The standard method is to isolate mMA from donor cells that highly express the gene and perform reverse transcription to form a plasmid or phage cDNA library. There are many mature techniques for extracting mRNA, and kits are also commercially available (Qiagene). The construction of cDNA libraries is also a common method (Sambrook, et al., Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory. New York, 1989). Commercially available cDNA libraries are also available, such as different cDNA libraries from Clontech. When polymerase reaction technology is used in combination, even very small expression products can be cloned.

 The genes of the present invention can be selected from these cDNA libraries by conventional methods. These methods include (but are not limited to): (l) DM-DM or DNA-RNA hybridization; (2) the presence or absence of marker gene functions; (3) measuring the level of human ribosomal protein L23 transcripts; (4) Detection of gene-expressed protein products by immunological techniques or determination of biological activity. The above methods can be used singly or in combination.

 In the method (1), the probe used for hybridization is homologous to any part of the polynucleotide of the present invention, and its length is at least 10 nucleotides, preferably at least 30 nucleotides, more preferably At least 50 nucleotides, preferably at least 100 nucleotides. In addition, the length of the probe is usually within 2,000 nucleotides, preferably within 1000 nucleotides. The probe used here is generally a DNA sequence chemically synthesized based on the gene sequence information of the present invention. The genes or fragments of the present invention can of course be used as probes. DNA probes can be labeled with radioisotopes, luciferin, or enzymes (such as alkaline phosphatase).

 In the method (4), the protein product of human ribosomal protein L23 gene expression can be detected by immunological techniques such as Western blotting, radioimmunoprecipitation, and enzyme-linked immunosorbent assay (ELISA).

 A method for amplifying DNA / RNA by PCR (Saiki, et al. Science 1985; 230: 1350-1354) is preferably used to obtain the gene of the present invention. In particular, when it is difficult to obtain full-length cDNA from a library, the RACE method (RACE-rapid amplification of cDNA ends) can be preferably used. The primers used for PCR can be appropriately based on the polynucleotide sequence information of the present invention disclosed herein. Select and synthesize using conventional methods. The amplified DNA / RNA fragments can be isolated and purified by conventional methods such as by gel electrophoresis.

 The polynucleotide sequence of the gene of the present invention or various DNA fragments and the like obtained as described above can be determined by a conventional method such as dideoxy chain termination method (Sanger et al. PNAS, 1977, 74: 5463-5467). Such polynucleotide sequences can also be determined using commercial sequencing kits. In order to obtain the full-length cDNA sequence, sequencing must be repeated. Sometimes it is necessary to determine the cDNA sequence of multiple clones in order to splice into a full-length cDNA sequence.

The present invention also relates to a vector comprising the polynucleotide of the present invention, and a host cell that is genetically engineered using the vector of the present invention or directly using the human ribosomal protein L23 coding sequence, and that the present invention is produced by recombinant technology. A method of inventing the polypeptide.

 In the present invention, a polynucleotide sequence encoding human ribosomal protein L23 can be inserted into a vector to constitute a recombinant vector containing the polynucleotide of the present invention. The term "vector" refers to bacterial plasmids, phages, yeast plasmids, plant cell viruses, mammalian cell viruses such as adenoviruses, retroviruses, or other vectors well known in the art. Vectors suitable for use in the present invention include, but are not limited to: T7 promoter-based expression vectors (Ros enberg, et al. Gene, 1987, 56: 125) expressed in bacteria; pMSXND expression vectors expressed in mammalian cells (Lee and Na thans, JBio Chem. 263: 3521, 1988) and baculovirus-derived vectors expressed in miracidial cells. In short, as long as it can be replicated and stabilized in the host, any plasmid and vector can be used to construct a recombinant expression vector. An important feature of expression vectors is that they usually contain replication origins, promoters, marker genes, and translational regulatory elements.

Methods well known to those skilled in the art can be used to construct expression vectors containing a DNA sequence encoding human ribosomal protein L23 and appropriate transcriptional / translational regulatory elements. These methods include in vitro recombination MA technology, DNA synthesis technology, in vivo recombination technology, etc. (Sambroook, et al. Mo lecul ar Cloning, a Labora tory Manua, cold Harbor Labora tory. New York, 1989). The DNA sequence 'J' can be effectively linked to an appropriate promoter in an expression vector to guide mRNA synthesis. Representative examples of these promoters are: the l ac or trp promoter of E. coli; the _PL promoter of lambda phage; eukaryotic promoters include the CMV immediate early promoter, the HSV thymidine kinase promoter, and the early and late SV40 promoters Promoters, retroviral LTRs, and other known promoters that control the expression of genes in prokaryotic or eukaryotic cells or their viruses. The expression vector also includes a ribosome binding site and a transcription terminator for translation initiation. Insertion of enhancer sequences into the vector will enhance its transcription in higher eukaryotic cells. Enhancers are cis-acting factors expressed by DM, usually about 10 to 300 base pairs, which act on promoters to enhance gene transcription. Illustrative examples include SV40 enhancers of 100 to 270 base pairs on the late side of the origin of replication, polyoma enhancers on the late side of the origin of replication, and adenovirus enhancers.

 In addition, the expression vector preferably contains one or more selectable marker genes to provide phenotypic traits for selection of transformed host cells, such as dihydrofolate reductase, neomycin resistance, and green for eukaryotic cell culture. Fluorescent protein (GFP), or tetracycline or ampicillin resistance for E. coli.

 Those of ordinary skill in the art will know how to select appropriate vector / transcription control elements (such as promoters, enhancers, etc.) and selectable marker genes.

In the present invention, a polynucleotide encoding human ribosomal protein L23 or a recombinant vector containing the polynucleotide Attach

It can be transduced or transduced into host cells to constitute a genetically engineered host cell containing the polynucleotide or recombinant vector. The term "host cell" refers to a prokaryotic cell, such as a bacterial cell; or a lower eukaryotic cell, such as yeast.

Cells; or higher eukaryotic cells, such as mammalian cells. Representative examples are: E. coli, Streptomyces; bacterial cells such as Salmonella typhimurium; fungal cells such as yeast; plant cells; insect cells such as flies

S2 or Sf 9; animal cells such as bright CH0, COS or Bowes melanoma cells.

Transformation of a host cell with a DNA sequence described in the present invention or a recombinant vector containing the DNA sequence can be performed using conventional techniques well known to those skilled in the art. When the host is a prokaryote such as E. coli, competent cells capable of absorbing DNA can be harvested after the exponential growth phase and treated with CaCl *. The steps used are well known in the art. The alternative is to use MgC l ₂ . If necessary, transformation can also be performed by electroporation. When the host is a eukaryotic organism, the following DNA transfection methods can be used: calcium phosphate co-precipitation method, or conventional mechanical methods such as microinjection, electroporation, and liposome packaging.

 Using conventional recombinant DNA technology, the polynucleotide sequence of the present invention can be used to express or produce recombinant human ribosomal protein L23 (Scence, 1984; 224: 1431). Generally there are the following steps:

 (1) using the polynucleotide (or variant) encoding human human ribosomal protein L23 of the present invention, or transforming or transducing a suitable host cell with a recombinant expression vector containing the polynucleotide;

 (2) culturing host cells in a suitable medium;

 (3) Isolate and purify protein from culture medium or cells.

 In step (2), the medium used in the culture may be selected from various conventional mediums according to the host cells used. Culture is performed under conditions suitable for host cell growth. After the host cells have grown to an appropriate cell density, the selected promoter is induced by a suitable method (such as temperature conversion or chemical induction), and the cells are cultured for a period of time.

In step (3), the recombinant polypeptide may be coated in a cell, expressed on a cell membrane, or secreted outside the cell. If desired, recombinant proteins can be isolated and purified by various separation methods using their physical, chemical, and other properties. These methods are well known to those skilled in the art. These methods include, but are not limited to: conventional renaturation treatment, protein precipitant treatment (salting out method), centrifugation, osmotic disruption, ultrasonic treatment, ultracentrifugation, molecular sieve chromatography (gel filtration), adsorption chromatography, ionization Exchange chromatography, high performance liquid chromatography (HPLC), and various other liquid chromatography techniques and combinations of these methods. The following drawings are used to illustrate specific embodiments of the invention, but not to limit the scope of the invention as defined by the claims.

 Fig. 1 is a comparison diagram of the amino acid sequence homology of the human ribosomal protein L23 and L22 proteins of the present invention. The upper sequence is the human ribosomal protein L23, and the lower sequence is the L22 protein. Identical amino acids are represented by single character amino acids between the two sequences, and similar amino acids are represented by "+".

 Figure 2 is a polyacrylamide gel electrophoresis image (SDS-PAGE) of the isolated human ribosomal protein L23. 23kDa is the molecular weight of the protein. The arrow indicates the isolated protein band. The best way to implement the invention

 The present invention is further described below with reference to specific embodiments. It should be understood that these examples are only used to illustrate the present invention and not to limit the scope of the present invention. The experimental methods without specific conditions in the following examples are usually performed according to the general conditions such as Sambrook et al., Molecular Cloning: Laboratory Manual (New York: Colling Harbor Laboratory Press, 1989), or following Conditions recommended by the manufacturer.

 Example 1: Cloning of human ribosomal protein L23

 Human fetal brain total RNA was extracted by one-step method with guanidine isothiocyanate / phenol / chloroform. Quik mRNA Isolat ion Ki t

(Qiegene product) Isolate poly (A) mRNA from total RNA. 2ug poly (A) mRNA was formed into cDNA by reverse transcription. The Smart cDNA cloning kit (purchased from Clontech) was used to insert the cDNA fragment into the multiple cloning site of pBSK (+) vector (Clontech) to transform DH5α. The bacteria formed a cDNA library. Dye terminate cycle reaction ion sequencing kit (Perkin-Elmer) and ABI 377 automatic sequencer (Perkin-Elmer) were used to determine the 5 'and y-terminal sequences of all clones. The determined cDNA sequence was compared with the existing public DNA sequence database (Genebank), and it was found that the cDNA sequence of one of the clones 1012E11 was new DNA. The inserted cDNA fragment contained in this clone was determined in both directions by synthesizing a series of primers. The results showed that the full-length cDNA contained in the 1012E11 clone was 707 bp (as shown in Seq ID NO: 1), and there was a 621 bp open reading frame (0RF) from 20 bp to 640 bp, encoding a new protein (such as Seq ID NO : As shown in 2). We named this clone PBS-1012E11 and the encoded protein was named human ribosomal protein L23.

Example 2: Homologous search of cDNA clones

The sequence of the human ribosomal protein L23 of the present invention and the protein sequence encoded by the same are used by the Mas t program (Basiclocal Alignment search tool) [Altschul, SF et al. J. Mol. Biol. 1990; 215: 403-10], homology search was performed in databases such as Genbank and Swissport. The gene with the highest homology to the human ribosomal protein L23 of the present invention is a known L22 protein, and its encoded protein has the accession number X84708 in Genbank. The protein homology results are shown in Figure 1. The two are highly homologous with 33% identity; 52% similarity. Example 3: Cloning of a gene encoding human ribosomal protein L23 by RT-PCR

 CDNA was synthesized using fetal brain total RNA as a template and oligo-dT as a primer for reverse transcription reaction. After purification with Qiagene's kit, the following primers were used for PCR amplification:

 Primerl: 5'- GGTAGCGGGAGGGCGAAAGATGGC —3, (SEQ ID NO: 3)

 Primer2: 5'- TCAATTTTTGTTTATTTTTAGAAA -3 '(SEQ ID NO: 4)

 Primerl is a forward sequence starting at lbp of the 5th end of SEQ ID NO: 1;

 Primer2 is the 3 'end reverse sequence in SEQ ID NO: 1.

Amplification reaction conditions: 50 mmol / L KC1, 10 legs ol / L Tris-CI, (pH8.5), 1.5 mmol / L MgCl ₂ , 200 μ mol / L dNTP, lOpmol in a 50 μ 1 reaction volume Primer, 1U of Taq DNA polymerase (Clontech). The reaction was performed on a PE9600 DNA thermal cycler (Perkin-Elmer) under the following conditions for 25 cycles: 94 ° C 30sec; 55. C 30sec; 72. C 2min. At-? 0, (3-& (^ is a positive control and template blank is a negative control. Amplification products were purified using QIAGEN's kit, and TA cloning kit was connected to the PCR vector (Invitrogen's product)). The DNA sequence analysis results show that the DNA sequence of the PCR product is exactly the same as the 1-707bp shown in SEQ ID NO: 1. Example 4: Northern blot analysis of human ribosomal protein L23 gene expression:

Total RNA extraction in one step [Anal. Biochem 1987, 162, 156-159] ₀ This method involves acid guanidinium thiocyanate-chloroform extraction. That is, the tissue was homogenized with 4M guanidinium isothiocyanate-25raM sodium citrate, 0.2M sodium acetate (pH4.0), and 1 volume of phenol and 1/5 volume of chloroform-isoamyl alcohol (49: 1 ), Mix and centrifuge. Aspirate the aqueous layer, add isopropanol (0.8 vol) and centrifuge the mixture to obtain RNA precipitate. The resulting RNA pellet was washed with 70% ethanol, dried and dissolved in water. Using 20 g of RNA, electrophoresis was performed on a 1.2% agarose gel containing 20 μM 3- (N-morpholino) propanesulfonic acid (pH 7.0)-5 mM sodium acetate-IfflM EDTA-2.2M formaldehyde. It was then transferred to a nitrocellulose membrane. _A - ³² PdATP was used to prepare ³² P-labeled DNA probes by random primers. DNA used The probe is the PCR amplified human ribosomal protein L23 coding region sequence (QObp to 640bp) shown in FIG. The P- labeled probes (about 2 xl0 ⁶ cpm / ml) and RNA was transferred to a nitrocellulose membrane overnight at 42 ° C in a hybridization solution, the solution comprising 50% formamide -25mM KH ₂ P0 ₄ ( pH7.4)-5 χ SSC- 5 χ Denhardt's solution and 200 μg / ml salmon sperm DNA. After hybridization, the filter was washed in 1 x SSC-0.1% SDS at 55 ° C for 30 min. Then, Phosphor Imager was used for analysis and quantification. Example 5: In vitro expression, isolation and purification of recombinant human ribosomal protein L23

 According to SEQ ID NO: 1 and the coding region sequence shown in FIG. 1, a pair of specific amplification primers was designed. The sequences are as follows:

 Primer3: 5'- CCCCATATGATGGCGGCGGCAGTACTGGGACAG —3, (Seq ID No: 5)

 Primer4: 5,-CATGGATCCTCATAGAGTGTGAACGATGGTCC -3, (Seq ID No: 6)

The 5 'ends of these two primers contain Ndel and BamHI restriction sites, respectively, followed by the coding sequences of the 5' and 3 'ends of the target gene, respectively. The Ndel and BamHI restriction sites correspond to the expression vector plasmid pET-28b ( +) (Novagen, Cat. No. 69865.3). PCR was performed using the pBS-1012Ell plasmid containing the full-length target gene as a template. The PCR reaction conditions were as follows: a total volume of 50 μl, containing 10 pg of pBS-1012E11 plasmid, primers Primer 3 and Primer-4, and j is lOpmol, Advantage polymerase Mix (Clontech) 1 μ1. Cycle parameters: 94 ° C 20s, 60 ° C 30s, 68. C 2 min, a total of 25 cycles. Ndel and BamHI were used to double-digest the amplified product and the plasmid PET-28 (+), respectively, and large fragments were recovered and ligated with T4 ligase. The ligation product was transformed into E. coli DH5CC using the calcium chloride method. After being cultured overnight on LB plates containing kanamycin (final concentration 30 g / ml), positive clones were screened by colony PCR method and sequenced. A positive clone (PET-1012E11) with a correct sequence was selected, and the recombinant plasmid was transformed into E. coli BL21 (DE3) plySs (product of Novagen) using the calcium chloride method. The host strain BL (pET-1012E11) was 37 in LB liquid medium containing kanamycin (final concentration 30 g / ml). C. Cultivate to logarithmic growth phase, add IPTG to a final concentration of 1 mmol / L, and continue to cultivate for ⁵ hours. The bacteria were collected by centrifugation, and the supernatant was collected by centrifugation. The supernatant was collected by centrifugation. The affinity chromatography column His. Bind Quick Cartridge (product of Novagen) was used to obtain 6 histidine (6His-Tag). The purified human ribosomal protein L23 was purified. After SDS-PAGE electrophoresis, a single band was obtained at 23 kDa (Figure 2). The band was transferred to a PVDF membrane and the N-terminal amino acid sequence was analyzed by the Edams hydrolysis method. As a result, the 15 amino acids at the N-terminus were identical to the 15 amino acid residues at the N-terminus shown in SEQ ID NO: 2. Example 6 Production of anti-human ribosomal protein L23 antibodies

 The following peptides specific to human ribosomal protein L23 were synthesized using a peptide synthesizer (product of PE):

NH ₂ -Met-A 1 a— A la— A la-Va l-Leu- G 1 y-Gln-Leu-G 1 yA 1 a-Leu-Trp-11 e- H i s-COOH (SEQ ID NO : 7). The polypeptide is coupled with hemocyanin and bovine serum albumin to form a complex, respectively. For the method, see: Avrameas, et al. Immunochemis try, 1969; 6: 43. Rabbits were immunized with 4 mg of the hemocyanin polypeptide complex plus complete Freund's adjuvant, and 15 days later, the hemocyanin polypeptide complex plus incomplete Freund's adjuvant was used to boost immunity once. A titer plate coated with a 15 g / ml bovine serum albumin peptide complex was used as an ELISA to determine antibody titers in rabbit serum. Total IgG was isolated from antibody-positive rabbit sera using protein A-Sepharose. The peptide was bound to a cyanogen bromide-activated Seph _arOS e4B column, and anti-peptide antibodies were separated from the total IgG by affinity chromatography. The immunoprecipitation method proved that the purified antibody could specifically bind to human ribosomal protein L23. Industrial applicability

 The polypeptide of the present invention and the antagonists, agonists and inhibitors of the polypeptide can be directly used in the treatment of diseases, for example, it can treat malignant tumors, adrenal deficiency, skin diseases, various inflammations, HIV infections and immune diseases.

Ribosome proteins play an important role in the process of ribosomal translation. Deletion or inappropriate chemical modification of ribosomal proteins, or mutations in ribosomal protein genes, will affect the function of ribosomes and reduce the activity of peptide synthesis. It leads to various serious consequences, such as unsustainable cell membrane structure, abnormal cell metabolism, cell growth arrest, and even death, etc., which causes various diseases, such as developmental disorders, metabolic diseases, immune diseases, cell death, and so on. Specifically in terms of the present invention, the polypeptide or fragment thereof of the present invention can be used to treat or prevent diseases caused by developmental disorders, including but not limited to: spina bifida, anencephaly, congenital hydrocephalus, aqueduct malformation , Cartilage hypoplasia, dwarfism, pseudochondral hypoplasia, gonad hypoplasia, etc. The polypeptide or fragment thereof of the present invention can also be used to treat or prevent diseases caused by metabolic disorders, including but not limited to: phenylketonuria , Albinism, trimethylamineuria, hypersarcosinemia, etc .; the polypeptides or fragments thereof of the present invention can also be used to treat or prevent various immunodeficiency disorders, including but not limited to: HIV, rheumatoid arthritis, chronic Active hepatitis, systemic lupus erythematosus, scleroderma, immune thrombocytopenic purpura, autoimmune interstitial nephritis, autoimmune heart disease, etc. The invention also provides screening compounds to identify raising (agonist) or repressing (antagonist) human ribosomal eggs. White L23 medicament method. Agonists enhance biological functions such as human ribosomal protein L23 to stimulate cell proliferation, while antagonists prevent and treat disorders related to excessive cell proliferation, such as various cancers. For example, mammalian cells or a membrane preparation expressing human ribosomal protein L23 can be cultured together with labeled human ribosomal protein L23 in the presence of a drug. The ability of the drug to increase or block this interaction is then determined.

 Antagonists of human ribosomal protein L23 include antibodies, compounds, receptor deletions, and analogs that have been screened. Antagonists of human ribosomal protein L23 can bind to human ribosomal protein L23 and eliminate its function, or inhibit the production of the polypeptide, or bind to the active site of the polypeptide so that the polypeptide cannot perform biological functions.

 When screening compounds as antagonists, human ribosomal protein L23 can be added to a bioanalytical assay to determine whether the compound is an antagonist by measuring the effect of the compound on the interaction between human ribosomal protein L23 and its receptor. Receptor deletions and analogs that act as antagonists can be screened in the same manner as described above for screening compounds. Polypeptide molecules capable of binding to human ribosomal protein L23 can be obtained by screening a random peptide library composed of various possible combinations of amino acids bound to a solid phase. When screening, the human ribosomal protein L23 molecule should generally be labeled.

 The present invention provides a method for producing an antibody using a polypeptide, a fragment, a derivative, an analog thereof, or a cell thereof as an antigen. These antibodies can be polyclonal or monoclonal antibodies. The invention also provides antibodies directed against the human ribosomal protein L23 epitope. These antibodies include (but are not limited to): polyclonal antibodies, monoclonal antibodies, 'chimeric antibodies, single chain antibodies, Fab fragments, and fragments produced by Fab expression libraries.

 Polyclonal antibodies can be produced by injecting human ribosomal protein L23 directly into immunized animals (such as rabbits, mice, rats, etc.). A variety of adjuvants can be used to enhance the immune response, including but not limited to Freund's adjuvant. . Techniques for preparing monoclonal antibodies against human ribosomal protein L23 include, but are not limited to, hybridoma technology (Kohler and Mistein. Nature, 1975, 256: 495-497), triple tumor technology, human beta-cell hybridoma technology, EBV-hybridoma technology, etc. Chimeric antibodies that bind human constant regions and non-human variable regions can be produced using existing techniques (Morrison et al., PNAS, 1985, 81: 6851). The existing technology for producing single chain antibodies (U.S. Pat No. 4946778) can also be used to produce single chain antibodies against human ribosomal protein L23.

 Antibodies against human ribosomal protein L23 can be used in immunohistochemistry to detect human ribosomal protein L23 in biopsy specimens.

Monoclonal antibodies that bind to human ribosomal protein L23 can also be labeled with radioisotopes and injected into the body to track their location and distribution. This radiolabeled antibody can be used as a non-invasive diagnostic method Localization of tumor cells and judgment of metastasis.

 Antibodies can also be used to design immunotoxins that target a particular part of the body. For example, human ribosomal protein L2 3 high affinity monoclonal antibodies can covalently bind to bacterial or plant toxins (such as diphtheria toxin, ricin, ormosine, etc.). A common method is to attack the amino group of an antibody with a thiol cross-linking agent such as SPDP and bind the toxin to the antibody through the exchange of disulfide bonds. This hybrid antibody can be used to kill human ribosomal protein L2 3 positive cells .

 The antibodies in the present invention can be used to treat or prevent diseases related to human ribosomal protein L23. Administration of an appropriate dose of antibody can stimulate or block the production or activity of human ribosomal protein L23.

 The invention also relates to a diagnostic test method for quantitatively and locally detecting the level of human ribosomal protein L23. These tests are well known in the art and include FI SH assays and radioimmunoassays. The level of human ribosomal protein L2 3 detected in the test can be used to explain the importance of human ribosomal protein L23 in various diseases and to diagnose diseases in which human ribosomal protein L23 plays a role.

 The polypeptide of the present invention can also be used for peptide mapping analysis. For example, the polypeptide can be specifically cleaved by physical, chemical or enzymatic analysis, and subjected to one-dimensional or two-dimensional or three-dimensional gel electrophoresis analysis, and more preferably mass spectrometry.

 The polynucleotide encoding human ribosomal protein L23 can also be used for a variety of therapeutic purposes. Gene therapy technology can be used to treat abnormal cell proliferation, development or metabolism caused by the non-expression or abnormal / inactive expression of human ribosomal protein L23. Recombinant gene therapy vectors (such as viral vectors) can be designed to express mutated human ribosomal protein L23 to inhibit endogenous human ribosomal protein L23 activity. For example, a mutated human ribosomal protein L23 may be a shortened human ribosomal protein L2 3 lacking a signaling domain, and although it can bind to downstream substrates, it lacks signaling activity. Therefore, the recombinant gene therapy vector can be used for treating diseases caused by abnormal expression or activity of human ribosomal protein L23. Virus-derived expression vectors such as retrovirus, adenovirus, adenovirus-associated virus, herpes simplex virus, parvovirus, etc. can be used to transfer the polynucleotide encoding human ribosomal protein L23 into cells. Methods for constructing recombinant viral vectors carrying a polynucleotide encoding the human ribosomal protein L23 can be found in the existing literature (Sambrook, et al.). In addition, a recombinant polynucleotide encoding human ribosomal protein L23 can be packaged into liposomes and transferred into cells.

 Methods for introducing a polynucleotide into a tissue or cell include: directly injecting the polynucleotide into a tissue in vivo; or introducing the polynucleotide into a cell in vitro through a vector (such as a virus, phage, or plasmid), and then transplanting the cell Into the body and so on.

Oligonucleotides (including antisense RNA and DNA) and ribozymes that inhibit human ribosomal protein L23 mRNA are also present Within the scope of the invention. A ribozyme is an enzyme-like RNA molecule that can specifically decompose specific RNA. Its mechanism of action is that the ribozyme molecule specifically hybridizes with a complementary target RNA and performs endonucleation. Antisense RNA and DNA and ribozymes can be obtained by any existing RNA or DNA synthesis technology, such as the technology for the synthesis of oligonucleotides by solid-phase phosphoramidite chemical synthesis has been widely used. Antisense RNA molecules can be obtained by in vitro or in vivo transcription of a DNA sequence encoding the RNA. This DNA sequence has been integrated downstream of the vector's RNA polymerase promoter. In order to increase the stability of a nucleic acid molecule, it can be modified in a variety of ways, such as increasing the sequence length on both sides, and the ribonucleoside linkages should use phosphate thioester or peptide bonds instead of phosphodiester bonds.

 The polynucleotide encoding human ribosomal protein L23 can be used for the diagnosis of diseases related to human ribosomal protein L23. The polynucleotide encoding human ribosomal protein L23 can be used to detect the expression of human ribosomal protein L2 3 or the abnormal expression of human ribosomal protein L23 in a disease state. For example, the DNA sequence encoding human ribosomal protein L23 can be used to hybridize biopsy specimens to determine the expression of human ribosomal protein L23. Hybridization techniques include Southern blotting, Nor thern blotting, and in situ hybridization. These technical methods are all mature technologies that are publicly available, and related kits are commercially available. Part or all of the polynucleotides of the present invention can be used as probes to be fixed on a microarray (Microcroix) or a DM chip (also known as a "gene chip") for analyzing differential expression analysis of genes in tissues and genes diagnosis. Human ribosomal protein L2 3 specific primers for RNA-polymerase chain reaction (RT-PCR) in vitro amplification can also detect the human ribosomal protein L23 transcription product.

 Detection of mutations in the human ribosomal protein L23 gene can also be used to diagnose human ribosomal protein L2 3 related diseases. Human ribosomal protein L23 mutations include point mutations, translocations, deletions, recombinations, and any other abnormalities compared to the normal wild-type human ribosomal protein L2 3 DNA sequence. Mutations can be detected using existing techniques such as Sou thern blotting, DNA sequence analysis, PCR and in situ hybridization. In addition, mutations may affect protein expression. Therefore, Nor thern blotting and Western blotting can be used to indirectly determine whether a gene is mutated.

 The sequences of the invention are also valuable for chromosome identification. This sequence will specifically target a specific position of a human chromosome and can hybridize with it. Currently, specific sites for each gene on the chromosome need to be identified. Currently, only a few chromosome markers based on actual sequence data (repeat polymorphisms) are available for labeling chromosomal positions. According to the present invention, in order to associate these sequences with disease-related genes, an important first step is to locate these DNA sequences on a chromosome.

In short, a PCR primer (preferably 15-35b _P ) is prepared from the cDNA, and the sequence can be mapped on the chromosome. These primers were then used for PCR screening of somatic hybrid cells containing individual human chromosomes. Only that Hybrid cells containing human genes corresponding to the primers will produce amplified fragments.

 PCR localization of somatic hybrid cells is a quick way to localize DNA to specific chromosomes. Using the oligonucleotide primers of the present invention, by a similar method, a set of fragments from a specific chromosome or a large number of genomic clones can be used to achieve sublocalization. Other similar strategies that can be used for chromosomal localization include in situ hybridization, chromosome pre-screening with labeled flow sorting, and pre-selection of hybridization to construct chromosome-specific cDNA libraries.

 Fluorescent in situ hybridization (FI SH) of cDNA clones with metaphase chromosomes allows precise chromosomal localization in one step. For a review of this technology, see Verma et al., Human Chromosomes: a Manu l of Basic Techniques, Pergamon Pres s, New York (1988).

 Once the sequence is located at the exact chromosomal location, the physical location of the sequence on the chromosome can be correlated with the genetic map data. These data can be found in, for example, V. Mckus ck, Mende l i an Inher i tance in Man (available online with Johns Hopkins University Welch Med ica l L brary). Linkage analysis can then be used to determine the relationship between genes and diseases that have been mapped to chromosomal regions.

 Next, the difference in cDM or genomic sequence between the affected and unaffected individuals needs to be determined. If a mutation is observed in some or all of the affected individuals and the mutation is not observed in any normal individual, the mutation may be the cause of the disease. Comparing affected and unaffected individuals usually involves first looking for structural changes in the chromosome, such as deletions or translocations that are visible at the chromosomal level or detectable with cDNA sequence-based PCR. According to the resolution capabilities of current physical mapping and gene mapping technology, the cDNA accurately mapped to the chromosomal region associated with the disease can be one of 50 to 500 potentially pathogenic genes (assuming 1 megabase mapping resolution) Capacity and each 20kb corresponds to a gene).

 The polypeptides, polynucleotides and mimetics, agonists, antagonists and inhibitors of the present invention can be used in combination with a suitable pharmaceutical carrier. These carriers can be water, glucose, ethanol, salts, buffers, glycerol, and combinations thereof. The composition comprises a safe and effective amount of the polypeptide or antagonist, and carriers and excipients that do not affect the effect of the drug. These compositions can be used as drugs for the treatment of diseases.

 The present invention also provides a kit or kit containing one or more containers containing one or more ingredients of the pharmaceutical composition of the present invention. Along with these containers, there may be instructional instructions given by government agencies that manufacture, use, or sell pharmaceuticals or biological products, which reminders permit their administration on the human body by government agencies that manufacture, use, or sell them. In addition, the polypeptide of the present invention can be used in combination with other therapeutic compounds.

The pharmaceutical composition can be administered in a convenient manner, such as by topical, intravenous, intraperitoneal, intramuscular, Subcutaneous, intranasal or intradermal route of administration. Human ribosomal protein L23 is administered in an amount effective to treat and / or prevent a specific indication. The amount and dose range of human ribosomal protein L23 administered to a patient will depend on many factors, such as the mode of administration, the health conditions of the person to be treated, and the judgment of the diagnostician.

(1) General information:

 (i i) Name of the invention: Human ribosomal protein L23 and its coding sequence

 (i i i) Number of sequences: 7

 (2) Information of SEQ ID D NO: 1:

 (i) Sequence characteristics:

(A) Length: 707b _P

 (B) Type: Nucleic acid

 (C) Chain: double strand

 (D) Topological structure: linear

 (i i) Molecular type: cDNA

 (x i) Sequence description: SEQ I D NO: 1:

GGTAGCGGGAGGGCGAAAGATGGCGGCGGCAGTACTGGGACAGTTGGGTGCGTTATGGAT ACATAACCTGAGGAGCCGGGGGAAGCTGGCCTTGGGTGTTTTACCTCAATCATATATCCA CACAAGTGCTTCTCTTGACATTTCTCGAAAATGGGAGAAGAAGAATAAAATTGTTTATCC TCCACAACTGCCTGGAGAACCTCGGAGACCAGCAGAAATCTACCACTGTCGAAGACAAAT AA AATATAGCAAAGACAAGATGTGGTATTTGGCAAA ATTG ATACGAGGAATGTCTATTGA CCAGGCTTTGGCTCAGTTGGAATTCA ATG ACA A AA AAGGGGCCA AA ATAATTAAAGAGGT TCTCTTAGAAGCACA AGATATGGCAGTG AGAGACCATAACGTGG AATTCAGGTCCAATTT ATATATAGCTGAGTCCACCTCAGGACGAGGCCAGTGCCTGAAACGCATCCGCTACCATGG CAGAGGTCGCTTTGGGATCATGGAGAAGGTTTATTGCCATTATTTTGTGA AGTTGGTGGA AGGGCCCCCACCTCCACCTGAGCCACCAAAGACGGCAGTTGCCCATGCCAAAGAGTATAT TCAGCAGCTTCGCAGCCGGACCATCGTTCACACTCTATGATGAGGAGATTCAGACTCCAC AGTGTATATATTTTGCCATTTATTTTCTAAAAATAAACAAAA ATTGA

(3) Information of SEQ ID NO: 2:

 (i) Sequence characteristics:

 (A) Length: 206 amino acids

 (B) Type: Amino acid

(D) Topological structure: linear (ii) Molecular type: peptide

 (xi) Sequence description: SEQ ID NO: 2:

 Met Ala Ala Ala Val Leu Gly Gin Leu Gly Ala Leu Trp lie His

Asn Leu Arg Ser Arg Gly Lys Leu Ala Leu Gly Val Leu Pro Gin

Ser Tyr lie His Thr Ser Ala Ser Leu Asp lie Ser Arg Lys Trp

Glu Lys Lys Asn Lys lie Val Tyr Pro Pro Gin Leu Pro Gly Glu

Pro Arg Arg Pro Ala Glu lie Tyr His Cys Arg Arg Gin lie Lys

Tyr Ser Lys Asp Lys Met Trp Tyr Leu Ala Lys Leu lie Arg Gly

Met Ser lie Asp Gin Ala Leu Ala Gin Leu Glu Phe Asn Asp Lys

Lys Gly Ala Lys lie lie Lys Glu Val Leu Leu Glu Ala Gin Asp

Met Ala Val Arg Asp His Asn Val Glu Phe Arg Ser Asn Leu Tyr lie Ala Glu Ser Thr Ser Gly Arg Gly Gin Cys Leu Lys Arg He

Arg Tyr His Gly Arg Gly Arg Phe Gly lie Met Glu Lys Val Tyr

Cys His Tyr Phe Val Lys Leu Val Glu Gly Pro Pro Pro Pro Pro

Glu Pro Pro Lys Thr Ala Val Ala His Ala Lys Glu Tyr He Gin

Gin Leu Arg Ser Arg Thr He Val His Thr Leu

(4) Information of SEQ ID NO: 3

 (i) Sequence characteristics

 (A) Length: 24 bases

 (B) Type: Nucleic acid

 (C) Chain: single chain

 (D) Topological structure: linear

 (Π) Molecular type: Oligonucleotide

 (xi) Sequence description: SEQ ID NO: 3:

GGTAGCGGGAGGGCGAAAGATGGC

(5) Information of SEQ ID NO: 4

 (i) Sequence characteristics

(A) Length: 24 bases (B) Type: Nucleic acid

(C) Chain: single chain

 (D) Topological structure: linear

 (i i) Molecular type: Oligonucleotide

 (x i) Sequence description: SEQ ID NO: 4:

TCAATTTTTGTTTATTTTTAGAAA

(6) Information of SEQ ID NO: 5

 (i) Sequence characteristics

 (A) Length: 32 bases

 (B) Type: Nucleic acid

 (C) Chain: single chain

 (D) Topological structure: linear

 (i i) Molecular type: Oligonucleotide

 (xi) Sequence description: SEQ ID NO: 5: CAGCCATGGCGGGGAAGAAGAATGTTCTGTCG

(7) Information of SEQ ID NO: 6

 (i) Sequence characteristics

 (A) Length: 29 bases

 (B) Type: Nucleic acid

 (C) Chain: single chain

 (D) Topological structure: linear

 (i i) molecular type: oligonucleotide

 (xi) Sequence description: SEQ ID NO: 6:

CCCGGATCCCGCTGCTTGGCCTTCTTCAC

(8) Information of SEQ ID NO: 7:

(i) Sequence characteristics: (A) Length: 15 amino acids

 (B) Type: Amino acid

 (D) Topological structure: linear

 (Π) Molecular type: Polypeptide

(xi) Sequence description: SEQ ID NO: 7:

 Met-Ala-Gly-Lys-Lys-Asn-Val-Leu-Ser-Ser-Leu-Ala-Val-Tyr-Ala

Claims

1. An isolated polypeptide-human ribosomal protein L23, characterized in that it comprises: a polypeptide having the amino acid sequence shown in SEQ ID NO: 2, or an active fragment, analog, or derivative thereof.

 2. The polypeptide according to claim 1, characterized in that the amino acid sequence of the polypeptide, analog or derivative has at least 95% identity with the amino acid sequence shown in SEQ ID NO: 2.

 3. The polypeptide according to claim 2, further comprising a polypeptide having an amino acid sequence represented by SEQ ID NO: 2.

 4. An isolated polynucleotide, characterized in that said polynucleotide comprises one selected from the group consisting of:

(a) a polynucleotide encoding a polypeptide having the amino acid sequence shown in SEQ ID NO: 2 or a fragment, analog, or derivative thereof;

 (b) a polynucleotide complementary to polynucleotide (a); or

 (c) A polynucleotide that is at least 70% identical to (a) or (b).

 5. The polynucleotide according to claim 4, wherein the polynucleotide comprises a polynucleotide encoding an amino acid sequence represented by SEQ ID NO: 2.

 6. The polynucleotide according to claim 4, characterized in that the sequence of the polynucleotide comprises a sequence of positions 20 to 640 in SEQ ID NO: 1 or a sequence of positions 1 to 707 in SEQ ID NO: 1. .

 7. A recombination vector containing an exogenous polynucleotide, characterized in that it is a recombination constructed by the polynucleotide according to any one of claims 4-6 and a plasmid, virus or a carrier expression vector Carrier.

 8. A genetically engineered host cell containing an exogenous polynucleotide, characterized in that it is selected from one of the following host cells:

 (a) a host cell transformed or transduced with the recombinant vector of claim 7; or

 (b) a host cell transformed or transduced with a polynucleotide according to any one of claims 4-6.

9. A method for preparing a polypeptide having human ribosomal protein L23 activity, characterized in that the method includes:

 (a) culturing the engineered host cell of claim 8 under the condition of expressing human ribosomal protein U3;

(b) Isolating a polypeptide with human ribosomal protein L23 activity from the culture.

10. An antibody capable of binding to a polypeptide, characterized in that the antibody is capable of binding to human ribosomal protein L23 Specific binding antibody.

 11. A class of compounds that mimic or regulate the activity or expression of a polypeptide, characterized in that they are compounds that mimic, promote, antagonize or inhibit the activity of human ribosomal protein L23.

 12. The compound according to claim 11, characterized in that it is an antisense sequence of a polynucleotide sequence or a fragment thereof as shown in SEQ ID NO: 1.

 13. Use of the compound according to claim 11, characterized in that the compound is used for a method for regulating the activity of human ribosomal protein L23 in vivo and in vitro.

 14. A method for detecting a disease or susceptibility to a disease associated with a polypeptide according to any one of claims 1-3, characterized in that it comprises detecting the expression level of the polypeptide, or detecting the activity of the polypeptide Or detecting a nucleotide variation in a polynucleotide that causes abnormal expression or activity of the polypeptide.

 15. Use of a polypeptide according to any one of claims 1-3, characterized in that it is used for screening mimetics, agonists, antagonists or inhibitors of human ribosomal protein L23; or for peptide fingerprinting Atlas identification.

 16. The use of a nucleic acid molecule according to any one of claims 4-6, characterized in that it is used as a primer for a nucleic acid amplification reaction, or as a probe for a hybridization reaction, or for manufacturing a gene chip Or microarray.

 17. Use of a polypeptide, polynucleotide or compound according to any one of claims 1-6 and 11, characterized in that the polypeptide, polynucleotide or mimetic, agonist, antagonist is used Or the inhibitor is composed of a safe and effective dose with a pharmaceutically acceptable carrier as a pharmaceutical composition for diagnosing or treating a disease associated with abnormality of human ribosomal protein L23.

 18. Use of a polypeptide, polynucleotide or compound according to any one of claims 1-6 and 11, characterized in that said polypeptide, polynucleotide or compound is used for preparing for treating malignant tumors, blood, etc. Disease, HIV infection and immune diseases and drugs of various inflammations.