Method for improving organic selenium synthesis ability of microorganisms based on sulfur-containing protein overexpression, and application thereof

A microorganism and organic selenium technology, applied in the field of selenium-enriched and organic selenium production, can solve the problems of exceeding the consumption capacity of ordinary people, high price of selenium-enriched yeast products, harsh nutritional conditions of brewer's yeast, etc., and achieve obvious and improved organic selenium conversion effect. Biomass yield and the effect of improving organic selenium yield

Active Publication Date: 2019-02-01
XI AN JIAOTONG UNIV
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AI Technical Summary

Problems solved by technology

[0006] At present, the main method to improve the selenium-enrichment ability of microorganisms is to obtain starting strains with higher selenium tolerance through artificial screening, and to increase the inorganic selenium content in the medium during the fermentation process so that microorganisms can enrich more selenoproteins to improve microbial growth. The organic selenium content in the body, but so far only found that brewer's yeast has a high ability to tolerate selenium, which may be related to its original high protein content
However, the nutritional conditions of brewer's yeast are harsh, and the yield when used in the production of selenium-enriched yeast is low, resulting in high production costs. The price of selenium-enriched yeast products is very high, exceeding the consumption capacity of ordinary people

Method used

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  • Method for improving organic selenium synthesis ability of microorganisms based on sulfur-containing protein overexpression, and application thereof
  • Method for improving organic selenium synthesis ability of microorganisms based on sulfur-containing protein overexpression, and application thereof
  • Method for improving organic selenium synthesis ability of microorganisms based on sulfur-containing protein overexpression, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Construction of artificially designed AC sulfur-containing protein recombinant yeast and determination of selenium-enriching ability

[0042] 1. Design of artificial sulfur-containing protein AC and its gene sequence

[0043] 1) Artificially design an alanine-cysteine ​​polymer as a sulfur-containing protein, with a total of 40 amino acids (AC40 for short), and its amino acid sequence is (AC)20. Among them, A represents alanine, C represents cysteine, and there are 20 sulfur-containing amino acids, accounting for 50%.

[0044] The total length of the AC gene sequence plus the start and stop codons is 126bp, and the sequence is as follows:

[0045] ATG (GCCTGC) 20 TGA (shown in SEQ.ID.NO.1).

[0046] 2) According to the yeast codon preference, optimize the codon sequence of AC40, determine its gene sequence, and add enzyme cutting sites upstream and downstream as follows:

[0047]

[0048] 2. Construction of recombinant expression vector

[0049] The ge...

Embodiment 2

[0064] Example 2 Construction of artificially designed MC sulfur-containing protein recombinant yeast and determination of selenium-enriching ability

[0065] 1. Design of artificial sulfur-containing protein MC and its gene sequence

[0066] 1) Artificially design a sulfur-containing protein composed of methionine and cysteine, with a total of 74 amino acids (referred to as MC74), which contains several gaps of glycine to meet the requirements of the protein structure, and the sequence is GG(MMMMMMGGCCGG)6. Among them, G represents glycine, M represents methionine, C represents cysteine, and the number of sulfur-containing amino acids is 48, accounting for about 65%. The total length of the MC gene sequence plus the start and stop codons is 228bp, and the sequence is as follows:

[0067] ATGGGTGGT (ATGATGATGATGATGATGGGTGGTTGTTGTGGTGGT) 6 T GA (as shown in SEQ.ID.NO.2).

[0068] 2) According to the yeast codon preference, optimize the codon sequence of MC74, determine its g...

Embodiment 3

[0085] Example 3 Construction of natural MT sulfur-containing protein recombinant bacteria and determination of selenium-enriching ability

[0086] 1. Natural rabbit metallothionein MT and its gene sequence

[0087] The most representative of natural sulfur-containing proteins is metallothionein MT, which is a metal-binding protein naturally formed in the process of biological evolution, and contains more sulfur-containing amino acids—cysteine. Studies have shown that the MT sequence is highly conserved, and the content of cysteine ​​is about 1 / 3 of the total amino acids. In this embodiment, the rabbit source MT protein is selected from the NCBI protein sequence database as a representative of natural sulfur-containing protein, and its amino acid sequence is:

[0088] MDPNCSCPTGGSCSCAGSCTCKACRCPSCKKSCCSCCPVGCAKCAQGCVCKGASDKCSCCA.

[0089] A total of 61 amino acids (MT61), of which C represents cysteine, a total of 20, sulfur-containing amino acids accounted for 32.8%. Addin...

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Abstract

The invention discloses a method for improving the organic selenium synthesis ability of microorganisms based on sulfur-containing protein overexpression , and belongs to the technical field of biosynthesis of organic selenium. An exogenous sulfur-containing protein gene is introduced into a microbial genome, and the obtained genome is cultured in a high-selenium and low-sulfur environment. Sulfurin sulfur-containing amino acids (cysteine and methionine) is replaced by selenium during the overexpression of an exogenous sulfur-containing protein to synthesize selenosulfate amino acid in order to synthesize a selenoprotein, so efficient organic selenium synthesis of the microorganisms is achieved. The method is based on a substitution competition relationship between selenium and sulfur,a process for synthesizing the organic selenium by using the selenoprotein is provided, the sulfur-containing foreign protein is constructed in a biotechnical way, and the selenoprotein that is the organic selenium is formed in transferred microorganisms. The method can efficiently enrich selenium and ensure the safety.

Description

technical field [0001] The invention belongs to the technical field of selenium enrichment, in particular to the technical field of organic selenium production, and relates to a method and application for improving microbial organic selenium synthesis ability / effect based on the overexpression of sulfur-containing proteins. Background technique [0002] Selenium is an indispensable nutrient element for the human body. In the human body, selenium can resist viruses, delay aging, regulate the human immune system, and also have various functions such as improving vision, improving memory, preventing rheumatic bone pain, breathing, and gastrointestinal diseases. Selenium deficiency can lead to cancer, diabetes, cardiovascular disease, and immune system disease. [0003] Compared with other essential trace elements for the human body, the distribution of selenium in the earth's crust is extremely uneven around the world, and high-selenium soils are often located in relatively sma...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/19C12N1/21C12N15/81C12N15/70C07K14/00A23L33/135A23L33/16A23K10/18A23K20/20C12R1/84C12R1/19
CPCA23V2002/00A23K10/18A23K20/30A23L33/135A23L33/16C07K14/00C12N15/70C12N15/815A23V2200/30A23V2250/1626
Inventor 杨水云赵浩淼廖丹李少峰徐文赵良瑞温玉荣高恩方庆梅
Owner XI AN JIAOTONG UNIV
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