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Method for improving catalytic activity of chloroperoxidase through chemical modification of small molecule fatty acid

A chloroperoxidase and catalytic activity technology, applied in the field of enzyme engineering, can solve problems such as difficulty, complex structure, and many influencing factors, and achieve the effect of improving high temperature thermal stability and improving catalytic activity

Inactive Publication Date: 2013-01-09
SHAANXI NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since enzyme protein is a biological macromolecule with a large molecular weight, complex structure, and many influencing factors, it is a very difficult challenge, and effective means are still relatively lacking; at the same time, the stability of enzymes that are ubiquitous at present and the requirements required for industrial production The contradiction between harsh conditions is also a bottleneck restricting the large-scale industrial application of enzymes

Method used

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  • Method for improving catalytic activity of chloroperoxidase through chemical modification of small molecule fatty acid
  • Method for improving catalytic activity of chloroperoxidase through chemical modification of small molecule fatty acid

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Embodiment 1

[0021] 1. Prepare the solution

[0022] Dissolve 0.142g octadecanoic acid in phosphate buffer solution with a mass fraction of 4% dimethyl sulfoxide to prepare 100mL of octadecanoic acid solution with a concentration of 0.5mmol / L; -4 The mol / L chloroperoxidase solution was diluted to 100 mL with phosphate buffer solution with a pH value of 5.0, and the concentration was prepared to be 5×10 -6 mol / L chloroperoxidase solution.

[0023] 2. Pretreatment dialysis bag

[0024] Boil the dialysis bag with 150 mL of 50% ethanol water solution by volume fraction for 1 hour, cool it down, and wash it with 50% ethanol water solution and 0.01mol / L NaHCO successively. 3 aqueous solution, 0.001mol / L ethylenediaminetetraacetic acid aqueous solution, washed with distilled water, then soaked in distilled water, and placed in a refrigerator at 4°C until use.

[0025] 3. Modification reaction

[0026] 3mL 5×10 -6 mol / L chloroperoxidase solution and 0.6mL 0.5mmol / L octadecanoic acid solution ...

Embodiment 2

[0030] In the preparation solution step 1 of embodiment 1, 0.1g dodecanoic acid is dissolved in the phosphate buffer solution of the dimethyl sulfoxide of 4% by mass fraction, preparation concentration is the dodecanoic acid solution 100mL of 0.5mmol / L, Other steps of this step are the same as in Example 1. In modification reaction step 3, 3 mL of 5×10 -6 mol / L chloroperoxidase solution and 0.6mL 0.5mmol / L dodecanoic acid solution were mixed evenly in a 5mL cryovial, placed in an ice bath and stirred for 60 minutes. The other steps were the same as in Example 1 to prepare dodecanoic acid-modified chloroperoxidase.

Embodiment 3

[0032] In the preparation solution step 1 of embodiment 1, 0.114g myristic acid is dissolved in the phosphate buffer solution of the dimethyl sulfoxide of 4% by mass fraction, and preparation concentration is 100mL of the myristic acid solution of 0.5mmol / L, Other steps of this step are the same as in Example 1. In modification reaction step 3, 3 mL of 5×10 -6 The mol / L chloroperoxidase solution and 0.6mL 0.5mmol / L myristic acid solution were mixed evenly in a 5mL cryovial, placed in an ice bath and stirred for 60 minutes. The other steps were the same as in Example 1 to prepare myristic acid-modified chloroperoxidase.

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Abstract

The invention relates to a method for improving catalytic activity of chloroperoxidase through chemical modification of small molecule fatty acid. In the method, the small molecule fatty acid such as dodecoic acid, tetradecanoic acid, octadecanoic acid, arachidic acid, behenic acid and the like are used for performing chemical modification on the chloroperoxidase, so the catalytic activity of thechloroperoxidase is effectively improved and high-temperature heat stability and anti-deactivation capacity for organic solvents are improved. The method is simple, and low in cost, and has great significance for industrialized application of the chloroperoxidase in the fields of organic synthesis and the like.

Description

technical field [0001] The invention belongs to the technical field of enzyme engineering, and in particular relates to a method for improving the catalytic activity of enzymes through chemical modification of small molecules. Background technique [0002] Enzyme-catalyzed reactions have been widely used in organic synthesis, pharmaceutical preparation and other fields due to their characteristics of mild conditions, high efficiency and orientation, and environmental friendliness. is of great significance. [0003] Chloroperoxidase (CPO) is a heme glycoproteinase isolated from the marine fungus Caldariomyces fumago. Due to its unique active site structure, CPO has both peroxidase, catalase and cytochrome P The catalytic function of -450 is currently considered to be the most widely used enzyme in the known heme peroxidase family, and has a wide range of substrate adaptability, so CPO is widely used in environmental biocatalysis, pharmaceutical industry and modern synthetic ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/96
Inventor 蒋育澄李超男胡满成翟全国李淑妮
Owner SHAANXI NORMAL UNIV