otud3-e86k protein and its coding gene and application

A protein and protein technology, applied in the fields of application, genetic engineering, plant gene improvement, etc., can solve the problem of undisclosed correlation between biological functions and diseases

Active Publication Date: 2016-09-21
INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] OTUD3 is an important protein identified for the first time in the world. Up to now, only the gene sequence, protein sequence and the spatial structure of its OTU domain have been disclosed in the world about OTUD3. Regarding its biological function and disease correlation, there is no no disclosure

Method used

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  • otud3-e86k protein and its coding gene and application
  • otud3-e86k protein and its coding gene and application
  • otud3-e86k protein and its coding gene and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1. Obtaining of overexpression OTUD3 lentivirus and knockdown OTUD3 expression lentivirus

[0062] 1. Obtaining of lentivirus overexpressing OTUD3

[0063] 1) Acquisition of OTUD3 protein and coding gene

[0064] The human OTUD3 gene contains 8 exons and 7 introns, the full length of its mRNA sequence is 6523nt (the nucleotide sequence is sequence 5), the CDS region is 1197nt (the nucleotide sequence is sequence 1), and it encodes a 398 amino acid protein OTUD3 (amino acid sequence is SEQ ID NO: 2).

[0065] OTUD3 protein mainly contains two structural domains: OTU (ovarian tumor domain) domain and UBA (ubiquitin-associated domain) domain. It was observed by confocal laser microscopy that OTUD3 was mainly localized in the cytoplasm of cells.

[0066] Using the purchased OTUD3 antibody, the expression of OTUD3 was detected in 12 breast cancer cell lines, HCT116 (colon cancer cells), HepG2 (liver cancer cells), HeLa (cervical cancer cells) and 293T (human embry...

Embodiment 2

[0092]Example 2, OTUD3 or the functional research of substances knocking down the expression of OTUD3

[0093] 1. The application of OTUD3 in inhibiting the proliferation of tumor cells

[0094] The lentivirus #1 which knocks down OTUD3 expression, the lentivirus which overexpresses OTUD3 and the negative control lentivirus prepared in Example 1 were used to infect breast cancer cell MCF7 (Peking Peking Union Medical College Cell Bank), and the knockdown was obtained by puromycin screening. Low OTUD3 expression cell line MCF7, OTUD3 overexpression cell line MCF7 and negative control cell line MCF7.

[0095] Will 1×10 3 The three above-mentioned cells were planted in 96-well plates respectively, and 0.05 mg ml -1 MTS (Promega), after further culturing at 37° C. for 4 hours, the absorbance value (OD490) at 490 nm light wave was detected. Calculate the corresponding cell number according to the standard curve.

[0096] The result is as figure 1 As shown in A:

[0097] The c...

Embodiment 3

[0219] Embodiment 3, discovery and acquisition of OTUD3 mutein

[0220] 1. Low expression of OTUD3 gene in tumor cells

[0221] Through the collection of clinical breast cancer patient samples and the purchase of commercial tumor tissue chips, more than 200 breast cancer samples and nearly 100 colon cancer samples have been accumulated.

[0222] The mRNA and protein levels of OTUD3 in cancer and paracancerous tissues of more than 60 breast cancer patients were detected (immunohistochemistry, antibodies from Abcam).

[0223] The result is as image 3 As shown in E, 3F and 3G, the mRNA and protein levels of OTUD3 were highly expressed in paracancerous tissues, but low in cancerous tissues.

[0224] Immunohistochemical analysis was performed on all cancer tissues, and the results were as follows image 3 H and 3I showed that OTUD3 was positively correlated with the expression of PTEN, a known important tumor suppressor factor.

[0225] It shows that the OTUD3 gene is lowly ex...

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Abstract

The invention discloses OTUD3-E86K protein as well as a coding gene and application thereof. The protein provided by the invention is (a) or (b) as follows: (a) protein consisting of amino acid sequences shown in a sequence 4 in a sequence table; (b) protein which is obtained by substitution, deletion and / or addition of one or more amino acid residues on the amino acid sequences shown in the sequence 4 in the sequence table, is same in function and is derived from the sequence 4. The experiment proves that the OTUD3-E86K protein can be used for designing or screening products for preventing and / or treating tumors, and provides important scientific basis and application values for tumor diagnosis, prevention and treatment.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an OTUD3-E86K protein, its coding gene and its application. Background technique [0002] The tumor suppressor protein PTEN (phosphatase and tension homologue deleted from chromosome 10, also known as MMAC1 or TEP1) gene is a new tumor suppressor gene located on human chromosome 10q23.3 by two independent research groups in the United States in 1997. The protein encoded by this gene, PTEN, is 403 amino acids long and consists of a phosphatase domain (15-185), a C2 domain (186-351), a PEST motif region (352-400) and a PDZ binding region (401-403). , PTEN has lipid phosphatase and protein phosphatase activities, more detailed research shows that its lipid phosphatase activity has a more important position in its tumor suppressor activity, it can specifically make phosphatidylinositol- The 3-phosphate group of 3,4,5-triphosphate (phosphatidylinositol-3,4,5-triphosphate, PIP3) is dephos...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/47C12N15/12C12N15/861C12N7/01C12N15/70C12N15/74C12Q1/68G01N33/68
CPCC07K14/47C12Q1/6886C12Q2600/158G01N33/6893G01N2800/50
Inventor 张令强贺福初袁林吕艳蓉李洪昌邢桂春
Owner INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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