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Study on the Expression and Purification of Caveolin-1 Protein and Its Relationship with Tumor

2021-04-30
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Caveolin exists in a variety of terminally differentiated cells such as fibroblasts, endothelial cells, and type I alveolar epithelium. It is a flask-shaped micro-recessed structure with a diameter of 50-100 nm on the cytoplasmic membrane. Caveolin-1 protein is the cellar Important structural protein. Studies have suggested that Caveolin-1 participates in the regulation of the activity of factors related to tumor occurrence, development, invasion and metastasis, and angiogenesis. The expression and purification of Caveolin-1 protein is conducive to obtaining a higher purity protein, which lays a foundation for further research on its biological activity.

Study on the expression and purification of Caveolin-1 protein

As an important part of natural life activities, proteins almost all exist in various structures of cells in a complex mixed form. It is very difficult to completely separate and purify the protein without destroying the spatial structure and primary structure of the protein without affecting its physiological function. In order to increase the purity of the protein sample and maintain the activity of the sample, protein separation and protein purification are required, and genetic engineering technology can be used to study protein expression and purification. Medicilon provides protein purification services. It has a variety of protein expression coordination, including prokaryotic protein expression system, yeast protein expression system, insect cell protein expression system (baculovirus), mammalian cell protein expression system, and a variety of fusion technologies. Can provide customers with a variety of options in protein expression and purification. From program design, gene optimization, expression condition optimization to purification technology system to improve the expression level of target protein.

Some researchers used genetic engineering methods to prepare the Caveolin-1 recombinant protein and its antibodies, laying the foundation for the study of the function of Caveolin-1 [1]. Methods The prokaryotic expression vector pGEX-5X-1-Caveolin-1 was constructed. After restriction enzyme digestion and identification, it was transformed into E. coli BL21 (DE3) and induced by IPTG. The expressed product was purified by SDS-PAGE and Western blot to identify the correctness of the gene fragment and the specificity of the expressed protein. After immunizing Balb/c mice with the purified recombinant protein 3 times, the titers of rabbit polyclonal antibodies were determined by ELISA.

As a result, the size of the cloned target gene fragment was consistent with expectations. The relative molecular weight of the recombinant Caveolin-1 protein is about 48,000, and the N-terminal fusion GST tag of the Caveolin-1 protein exists in the form of inclusion bodies to facilitate purification and identification. The obtained high-purity Caveolin-1 fusion protein can produce specific high-titer antibodies (ELISA titer up to 1:10000) after immunizing animals. The study constructed a prokaryotic expression system that highly expressed Caveolin-1 protein. The prepared Caveolin-1 recombinant protein immunized animals and obtained specific and high-titer antibodies, which provided experimental materials and basis for studying the biological activity of Caveolin-1 gene. .

The relationship between Caveolin-1 and tumors

Caveolin-1 was originally thought to be a tumor suppressor gene. Caveolin-1 mRNA and protein are highly expressed in normal tissues, but in neurogenic cells, ovarian cancer, colon cancer, sarcoma, lung cancer, breast cancer and many other tumor cell lines Decrease in expression. Caveolin-1 can inhibit tumor cell invasion and metastasis. The re-expression of Caveolin-1 in the oncogene-transformed cell line can inhibit cell growth and reduce its tumorigenicity.

Some researchers have studied the immunohistochemical detection of cervical cancer tissue Caveolin-1 and its mechanism of action in the occurrence and development of cervical cancer [2]. The method is to detect the expression of Caveolin-1 by immunohistochemistry. The results showed that Caveolin-1 presents brown-yellow or tan reactant particles in the cell membrane and cytoplasm. There are certain differences in the expression of Caveolin-1 in different cervical tissues. The expression rate of the tissues adjacent to the cancer is the highest. The expression rate was the lowest. The expression rates of Caveolin-1 in cervical invasive carcinoma and stage patients were 88.52% and 57.50%, respectively (P<0.05), and the expression rates in high, medium, and poorly differentiated carcinomas were 94.59% and 70.00%, respectively. , 50.00% (P<0.05). The decreased expression of Caveolin-1 in cervical cancer suggests that the decreased expression may be involved in the occurrence and development of cervical cancer.

The main cause of tumors is the activation of proto-oncogenes and the inactivation of tumor suppressor genes, which causes cell proliferation and differentiation to lose control. Caveolin-1 is sometimes described as a tumor suppressor, and sometimes it is also believed to promote tumor development. Studies have found that Caveolin-1 can promote cancer or inhibit the occurrence of cancer according to the type and stage of the tumor. The Caveolin-1 protein is expressed and purified by genetic engineering technology to obtain enough Caveolin-1 protein and the follow-up study of its regulatory mechanism for tumors is used for the development of anti-tumor drugs targeting Caveolin-1 in the future. Important.

[1] Cloning and expression of Caveolin-1 and preparation of its antibody [J].

[2] Caveolin-1 expression in cervical cancer tissues and its mechanism of action in the occurrence and development of cervical cancer[J].

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