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SeMet-labeled Protein High-yield Expression Medium

2021-11-29
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Selenium has been recognized by the World Health Organization as an essential trace element for the human body. It has strong anti-tumor and immunity enhancement effects. Compared with inorganic selenium, organic selenium has low toxicity, high bioavailability, and strong pharmacological activity. Selenomethionine is a natural organic selenium product with very few sources. Therefore, preparation by chemical synthesis is particularly important. Today, when tumors are increasingly threatening to human health, the use of selenomethionine for prevention and treatment is of great significance. In the United States, selenomethionine has been widely used as a medicine for cancer prevention and treatment. Adding selenium to some beverages and foods as a trace element can be used to improve the body’s immunity and prevent various selenium deficiency diseases. This product can be developed into medicines, health products, food additives and animal feed additives.

In addition, MAD (Multi-wavelength anomalous dispersion) is often used in crystallography to estimate the phase. Compared with sulfur atoms, the anomalous scattering signal of heavy atom selenium is clearer. Therefore, labeling proteins with selenomethionine is more and more widely used in MAD phase analysis experiments.

selenium selenomethionine

Firstly, the chemical properties of selenium are very close to sulfur; secondly, selenomethionine can completely replace methionine without changing the properties of the substituted protein; secondly, most of the protein crystals with selenomethionine substituted for methionine are the same shape as the corresponding natural protein crystals Crystal. The process of culturing protein crystals of selenomethionine is divided into the following four steps: expression, cell growth, purification and crystallization.

There are three options for the expression of selenomethionine-replaced methionine protein in E. coli. The plasmid containing the target gene can be transformed into the methionine-deficient (met-) strain of Escherichia coli to produce the methionine-deficient (met-) strain of E. ) Strains, and recently commonly used pathways to block methionine synthesis. The principle of the third method is that in Escherichia coli, high concentrations of isoleucine, lysine and threonine can block the synthesis of methionine by inhibiting the activity of aspartate kinase, and phenylalanine Acid and leucine act synergistically with lysine. Therefore, non-auxotrophic strains can contain sufficient selenomethionine and amino acids known to inhibit methionine synthesis without methionine.

Except for slightly stronger hydrophobicity and poorer solubility, the purified selenomethionine protein has properties similar to natural protein. To avoid oxidation of selenomethionine to selenium oxide, all buffers should be degassed, and DTT (5-20mM) and a chelating agent such as EDTA (0.2mM) should be added to remove metal ions that may contribute to oxidation. Minimize the purification time, such as using the FPLC system.
The crystallization conditions of selenomethionine-containing protein are similar to those of natural protein. Usually the protein containing selenomethionine is slightly less soluble, so the concentration of the protein or precipitant should be reduced accordingly. In addition to proteins that need to form disulfide bonds, DTT and EDTA are generally required for crystal culture of proteins containing selenomethionine.

Selenium Amino Acid Medium

Medicilon Biotech Services provides a full set of M9 selenomethionine (SeMET) medium, which can be used in IPTG-induced E. coli expression systems to produce selenomethionine-labeled proteins, and uses multi-wavelength anomalous scattering (MAD) methods for protein crystallography studies . This medium simplifies the steps of E. coli cultivation and saves you a lot of time, labor and material costs. After careful optimization by Medicilon, the Structural Genomics Laboratory can accelerate protein production; we can also specifically design and optimize the medium according to your needs. The medium does not need to be sterilized and weighs various components, just add one by one according to the enclosed manual. M9 high-yield medium has doubled the yield of bacteria, and it is more and more widely used in scientific research and biomedical industry.

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