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A kind of fungal laccase mutant pie5 and its expression strain and application

A fungal laccase and mutant technology, applied in the direction of fungi, enzymes, redox enzymes, etc., can solve the problems of inactivity, unfavorable treatment, environmental protection, and increased washing costs

Active Publication Date: 2021-08-31
ANHUI UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Since fungal laccase can only exert its optimal activity under acidic conditions (pH value 3-5), and under neutral or alkaline conditions, although fungal laccase is relatively stable, it has almost no activity
Therefore, in the finishing and dyeing process of denim, it is necessary to control the pH value of the washing at about 5, and after the reaction, the pH of the wastewater needs to be adjusted to neutrality. This process increases the cost of washing and increases the cost of washing. The concentration of salt ions in water is not conducive to post-processing and environmental protection

Method used

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  • A kind of fungal laccase mutant pie5 and its expression strain and application
  • A kind of fungal laccase mutant pie5 and its expression strain and application
  • A kind of fungal laccase mutant pie5 and its expression strain and application

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Embodiment Construction

[0029]The terms used in the present invention, unless otherwise specified, generally have the meanings commonly understood by those skilled in the art. In the following examples, various processes and methods not described in detail are conventional methods well known in the art.

[0030] 1. Preparation of fungal laccase PIE5

[0031] Strains: E.coli JM109, P.pastoris GS115

[0032] Vector: pPIC9K

[0033] Medium:

[0034] YPD medium (1L): 20g Peptone, 10g Yeast Extract dissolved in 900mL water, after autoclaving, add 100mL 10×D, add 1.5% agar to the solid medium.

[0035] The solvent of the seed liquid medium is PBS buffer solution with a pH value of 6.0 and 0.1M, and the solute and its concentration are as follows: YeastExtract 10g / L, Tryptone 20g / L, YNB 13.4g / L, glycerol 10g / L, biotin 4×10 -4 g / L.

[0036] The solvent of the fermentation liquid medium is PBS buffer solution with a pH value of 6.0 and 0.1M, and the solute and its concentration are as follows: YNB13.4 g / ...

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Abstract

The invention discloses a fungal laccase mutant PIE5 and its expression strain and application, wherein the fungal laccase mutant PIE5 transforms Coprinus cinerea laccase Lcc9 by means of directed evolution, constructs a mutation library, combines three rounds of Screening method to obtain laccase mutants to improve application ability of laccase. The fungal laccase mutant PIE5 of the present invention has an optimum catalytic pH of 8.5 (guaiacol is the substrate), and can still exert good activity when the pH reaches 9.5, so the fungal laccase PIE5 has an alkaline environment by comparison. Application advantages broaden the application field of fungal laccase, which can be directly applied to textile finishing process instead of stone mill washing, and there is no need to adjust the pH of wastewater in the degradation of wastewater indigo, which is conducive to improving the textile finishing process of fungal laccase, reducing Operating costs and protecting the ecological environment.

Description

technical field [0001] The invention relates to a fungal laccase mutant PIE5 and its expression strain and application. Background technique [0002] Laccase is a copper-containing polyphenol oxidase that can catalyze the oxidation of various substrates such as phenols and aromatic amines. When some small molecular substances exist as mediators, laccase can further broaden its oxidized substrates. and at the same time O 2 reduced to water. Laccase is widely distributed in plants, animals and microorganisms. Among them, microorganisms are the main source species of laccase. In general, fungal laccases have a high redox potential (470-790mV) and a high specific activity (100-1,000 U / mg). In comparison, the redox potential of bacterial laccase is lower (340-490mV), and the specific activity is also lower than that of fungal laccase (0.1-10 U / mg). Therefore, fungal laccase has wider application value. [0003] Due to the broad substrate specificity of fungal laccases, they...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/02C12N1/16D06P5/13C12R1/84
CPCC12N9/0061C12Y110/03002D06P5/132
Inventor 肖亚中尹强方泽民张学成房伟
Owner ANHUI UNIVERSITY
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