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Method for expression of Aspergillus flavus urate oxidase and the special gene thereof

A technology of uric acid oxidase and Aspergillus flavus, which is applied in the field of expressing Aspergillus flavus uric acid oxidase, can solve the problems of no acute gout attack, increased cost and process, long Uox cycle, etc., and achieves low cost, short cycle and high expression level. Effect

Active Publication Date: 2008-07-30
INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE
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Problems solved by technology

However, previous chemotherapy drugs, whether they inhibit uric acid synthesis or promote uric acid excretion, are suspected of causing acute gouty arthritis, and are generally not used for the treatment of acute attacks of gout
[0008] Aspergillus flavus Uox expressed by Saccharomyces cerevisiae abroad has a long cycle and is expensive
Some domestic researchers use prokaryotic expression system to express recombinant Aspergillus flavus Uox fused with the His tag. Although the His tag can be removed by protease, its N-terminus is unnatural, leaving 4 amino acid residues, which increases the cost of purification and craft

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  • Method for expression of Aspergillus flavus urate oxidase and the special gene thereof
  • Method for expression of Aspergillus flavus urate oxidase and the special gene thereof
  • Method for expression of Aspergillus flavus urate oxidase and the special gene thereof

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Embodiment 1

[0030] Embodiment 1, the synthesis of the optimized Aspergillus flavus UOX gene and the expression of uric acid oxidase

[0031] DNA restriction endonuclease, T4 DNA ligase, Pyrobest DNA polymerase and pMD-T vector were purchased from TaKaRa Company.

[0032] E.coli plasmid was extracted by using Omega kit; for the operation method of plasmid construction, refer to Molecular Cloning.

[0033] 1. Synthesis of rUox gene and construction of expression vector

[0034] Referring to the Aspergillus flavus UOX gene sequence (sequence number: X61766) and literature published by GenBank, analyze the complex secondary structure that may affect protein translation in Escherichia coli, optimize the gene, and keep the amino acid sequence unchanged Site-directed mutation was performed to remove the complex secondary structure. The mutated gene sequence was synthesized by Boya Biological Company and cloned into the pMD-T vector for sequence determination. DNA sequence determination confirme...

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Abstract

The invention discloses a method and special expression gene to express aspergillus flavus urate oxidase that contains one sequence from DNA sequence of SEQ ID No:3 in sequence table, and DNA sequence cross breeding nucleotide sequence of DNA sequence restricted by SEQ ID No:3 in sequence table. The invention has high expression level and could reach to 40% of solubility full mycoprotein, and is short cycle and low cost.

Description

technical field [0001] The invention relates to a method for expressing Aspergillus flavus uric acid oxidase and a special gene thereof. Background technique [0002] Uric acid oxidase (EC1.7.3.3, Uricase, Urate Oxidase, Uox) is an enzyme in the purine degradation and metabolism pathway in organisms, which catalyzes the oxidation of uric acid to generate allantoin, which is a metabolite that is relatively easy to excrete , its solubility is 5 to 10 times that of uric acid. Most mammals have endogenous Uox, but higher mammals (apes and humans) lack this enzyme and uric acid is excreted with urine as the end product. [0003] In normal human blood circulation, more than 99% of uric acid exists in the form of sodium urate (urate for short), and serum urate fluctuates within a narrow range. According to domestic data, the average male is 5.7mg / dl, and the average female is 4.3mg / dl. dl. If the concentration of uric acid in the serum is higher than the normal value, hyperurice...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/53C12N15/63C12N15/70C12N1/21C12N9/02
Inventor 陈薇李建民侯利华付玲于长明
Owner INST OF BIOENG ACAD OF MILITARY MEDICAL SCI OF THE CHINESE