Aspergillus niger inulin endopeptidase gene and recombinant Pichia strain for expressing same

A technology of inulin endoenzyme and Aspergillus niger strain, applied in genetic engineering, plant gene improvement, recombinant DNA technology, etc., can solve the problems of undetectable, low yield of enzyme protein, inconvenient industrial production, etc.

Inactive Publication Date: 2005-03-16
FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES +1
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Problems solved by technology

[0010] Using E.coli as the host bacteria, the expression research shows that the protein yield of inuA expression enzyme is very low and can

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  • Aspergillus niger inulin endopeptidase gene and recombinant Pichia strain for expressing same
  • Aspergillus niger inulin endopeptidase gene and recombinant Pichia strain for expressing same
  • Aspergillus niger inulin endopeptidase gene and recombinant Pichia strain for expressing same

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Embodiment Construction

[0014] The invention provides an Aspergillus nige strain (Aspergillus nige) 9891 producing high-yield endo-inulinase. The distribution of extracellular and intracellular inulinase activity measured with inulin as a substrate changes inversely with the culture time. The activity of extracellular enzymes decreased (Figure 1), and the activities of intracellular enzymes increased, and the absolute values ​​were all <4; However, it increased (Fig. 3 and Fig. 4), and the extracellular activity reached the highest activity unit <5.5, and the intracellular activity was as high as 50 IU.

[0015] The invention also provides a new inulin endonuclease gene. Cloning the inulin endonuclease gene inu9891 from producing inulin endonuclease Aspergillus niger strain 9891, it has an open reading frame encoding 495 amino acids, and this protein MW is 53.4KD; This gene and Aspergillus niger (Ohtak.et al, 1998) and Aspergillus ficuum (Uhm, T., et al., 1998) endinulinase gene homology is 94%; and...

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Abstract

The invention concerns aspergillus niger 9891 CGMCC NOú‘0991, which produces alantin endonuclease, and clones alantin endonuclease gene. The result of target gene fragment sequence analysis indicates that open reading frame of gene not containing signal peptide is 1485bp, and codes 509 amino acid, the said protein molecular weight is 55.9KD. The homolog of said gene with counterpart of aspergillus niger (Ohtak.et alú¼ 1998)and Aspergillus ficuum(Uhmú¼t.ú¼et alú¼1998) are respectively 92úÑ and 95úÑ. The alantin endonuclease gene is inserted into Pichia pastoris expression vector, thus recombinant of transformed Pichia pastoris is obtained. The said gene expresses in Pichia pastoris, and expressed product has its function. The expression amount of good recombinant I3-50 is 84 times higher than alantin endonuclease of initial strain. Recombinant yeast is induced and fermented. The analysis about recombinant enzyme shows that its optimum PH is 5.5ú¼ and optimum reaction temperature is 55íµ..

Description

technical field [0001] The present invention relates to the technical field of DNA, in particular, the present invention relates to Aspergillus niger strain Aspergillus niger 9891 producing endo-inulinase, and a new endo-inulinase gene cloned therefrom, and the present invention also relates to the gene containing said gene Recombinant Pichia pastoris, the expression level of the recombinant yeast endo-inulinase activity (using sucrose as substrate) is 84 times higher than that of the starting strain. Background technique [0002] Fructooligosaccharide (FOS) is an oligosaccharide compound composed of 3-10 fructose units. It is an effective regulator of microecological balance in animals and humans. It has been widely confirmed that oligosaccharides have a series of positive effects on the growth and development of humans and animals. effect. Including: 1) Promote animal weight gain, improve animal feed conversion efficiency, improve animal health, and increase daily weight ...

Claims

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Application Information

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IPC IPC(8): C12N1/14C12N1/19C12N9/24C12N15/56C12N15/81
Inventor 王建华滕达姚怡杨雅麟张帆
Owner FEED RESEARCH INSTITUTE CHINESE ACADEMY OF AGRICULTURAL SCIENCES
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