Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
2002-05-23
2004-11-02
Wax, Robert A. (Department: 1653)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C514S012200, C530S350000
Reexamination Certificate
active
06811987
ABSTRACT:
FIELD OF INVENTION
The invention relates to the field of biotechnology. In particular, the invention relates to a novel polypeptide, human calcium binding protein 42, and a polynucleotide sequence encoding said polypeptide. The invention also relates to the method for the preparation and use of said polynucleotide and polypeptide.
TECHNICAL BACKGROUND
Calcium functions as a medium in many biological activities in cell. These biological activities include gene regulation, cell cycle regulation, signal transduction, release of brain neurotransmitter and decomposition of glycogen involved in muscle contraction and so on. These functions are activated by calcium that are released by stimulation of extracellular signals. Calcium binds with various calcium binding protein to mediate further signals which activate other molecules and effect some specific physiological functions.
Calcium binding protein family is a very large protein super family with a calcium binding motif named “EF-hand,” which is the binding site of the protein and calcium. Many calcium binding proteins have several EF-hand motifs. Approximately 250 calcium binding proteins of this type have been identified.
Calmodulin (CaM) is the most widely distributed calcium regulatory protein. Calmodulin contains four EF-hand motifs. When they bind with calcium, corresponding conformation change will occur. The activation of calmodulin will cause it to bind with other target proteins and change their activities. The main target protein of calmodulin is calmodulin-dependent protein kinase, which is involved in the regulation of smooth muscle contraction, glycogen metabolism and neurotransmitter release, while calciuneurin, another calmodulin-binding protein, is involved in brain synaptic transmission. Calmodulin plays an important role in these physiological processes. Its abnormal expression often adversely affects these normal physiological processes and leads to corresponding diseases, such as those related to abnormal release of neurotransmitter, including nervous system development disorder, nervous system degenerative diseases, neuromuscular disorder, neurocutaneous syndrome and so on.
Calcyphosine is another calcium binding protein that is co-regulated by calcium binding and phosphorylation. The dog calcyphosine P24 is a calcium binding protein with 3 EF-hand motifs and phosphorylated by cAMP-dependent protein kinase (Lefort, A., et al. 1989, EMBO, 8: 111-116). P24 is expressed in various secretory tissues such as salivary gland, lung and brain. This shows that P24 plays an important role in ion transport. In 1993, Nemoto Y. et al cloned a similar calcium binding phosphorylating protein R2D2 that regulates the signal transduction in olfactory nerve cells from hare. R2D2 has 3 EF-hand motifs and is phophorylated by cAMP-dependent protein kinase and CaM kinase (Nemoto Y. et al., 1993, J. Cell Biol., 123: 963-76). In the same year, Yasuo Nemoto et al. cloned an R2D2 antigen protein of 189 amino acid from hare. This protein has 3 EF-hand motifs, a Ca2+/calmodulin-dependent protein kinase II phosphorylation site and a cAMP-dependent kinase phosphorylation site. It is the first calcium binding protein found in olfactory sensory neurons that regulate olfactory signal transduction (Yasuo Nemoto, Jun Ikeda et al., 1993, J. Cell Biol., 123: 963-976). It is involved in the olfactory function, and its abnormal expression will lead to diseases related to olfactory organs.
The human calcium binding protein 42 of the present invention shares 32% identity and 55% similarity with the hare R2D2 antigen. They possess similar structural characteristics, that is, both of them have calcium binding EF-hand motif, Ca2+/calmodulin-dependent protein kinaseII phosphorylation site and cAMP-dependent protein kinase phosphorylation site. So both of them are calcium binding protein.
DISCLOSURE OF INVENTION
One objective of the invention is to provide an isolated novel polypeptide, i.e., a human calcium binding protein 42, and fragments, analogues and derivatives thereof.
Another objective of the invention is to provide a polynucleotide encoding said polypeptide.
Another objective of the invention is to provide a recombinant vector containing a polynucleotide encoding a human calcium binding protein 42.
Another objective of the invention is to provide a genetically engineered host cell containing a polynucleotide encoding a human calcium binding protein 42.
Another objective of the invention is to provide a method for producing a human calcium binding protein 42.
Another objective of the invention is to provide an antibody against a human calcium binding protein 42 of the invention.
Another objective of the invention is to provide mimetics, antagonists, agonists, and inhibitors for the polypeptide of the human calcium binding protein 42.
Another objective of the invention is to provide a method for the diagnosis and treatment of the diseases associated with an abnormality of human calcium binding protein 42.
The present invention relates to an isolated polypeptide, which is originated from human, and comprises a polypeptide having the amino acid sequence of SEQ ID NO: 2, or its conservative variants, or its active fragments, or its active derivatives and its analogues. Preferably, the polypeptide has the amino acid sequence of SEQ ID NO: 2.
The present invention also relates to an isolated polynucleotide, comprising a nucleotide sequence or its variant selected from the group consisting of (a) the polynucleotide encoding a polypeptide comprising the amino acid sequence of SEQ ID NO: 2; and (b) a polynucleotide complementary to the polynucleotide (a); (c) a polynucleotide that shares at least 70% homology to the polynucleotide (a) or (b). Preferably, said nucleotide sequence is selected from the group consisting of (a) the sequence of position 1022-2170 in SEQ ID NO: 1; and (b) the sequence of position 1-3829 in SEQ ID NO: 1.
The invention also includes: a vector containing a polynucleotide of said invention, especially an expression vector; a host cell genetically engineered with the vector via transformation, transduction or transfection; a method for the production of said inventive polypeptide through the process of host cell cultivation and expression product harvest.
The invention also relates to an antibody which specifically binds to the inventive polypeptide.
The invention also relates to a method for selecting compounds which could simulate, activate, antagonize, or inhibit the activity of the inventive polypeptide and the compounds obtained by the method.
The invention also relates to a method for in vitro diagnosis method of the diseases or disease susceptibility related with the abnormal expression of the inventive polypeptide. The method involves the detection of mutation in the polypeptide or its encoding polynucleotide sequence, or the determination of its quantity and/or biological activity in biological samples.
The invention also relates to pharmaceutical compositions which comprises the inventive polypeptide, its analogues, mimetics, agonists, antagonists, inhibitors, and a pharmaceutically acceptable carrier.
The invention also relates to applications of the inventive polypeptide and/or its polynucleotide for drug development to treat cancers, developmental diseases, immune diseases, or other diseases caused by abnormal expression of the inventive polypeptide.
Other aspects of the invention are apparent to the skilled in the art in view of the disclosure set forth hereinbelow.
The terms used in this specification and claims have the following meanings, unless otherwise noted.
“Nucleotide sequence” refers to oligonucleotide, nucleotide, or polynucleotide and parts of polynucleotide. It also refers to genomic or synthetic DNA or RNA, which could be single stranded or double stranded, and could represent the sense strand or the antisense strand. Similarly, the term “amino acid sequence” refers to oligopeptide, peptide, polypeptide, or protein sequence and parts of proteins. When the “amino acid sequence” in the invention i
Mao Yumin
Xie Yi
Shanghai Bio Road Gene Development, Ltd.
Wax Robert A.
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