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Enzyme-metal ion nano compound and preparation method thereof

A nano-composite, metal ion technology, applied in biochemical equipment and methods, enzymes, oxidoreductases, etc., can solve problems such as enzyme inactivation and complex process, and achieve constant enzyme activity, simple operation, and good application. Foreground effect

Inactive Publication Date: 2016-07-13
TSINGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this method involves steps such as the synthesis of functionalized polyethylene glycol, the chemical modification of the enzyme, and the purification of the enzyme-polymer conjugate. The process is relatively complicated and easily causes the inactivation of the enzyme.

Method used

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  • Enzyme-metal ion nano compound and preparation method thereof
  • Enzyme-metal ion nano compound and preparation method thereof
  • Enzyme-metal ion nano compound and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Embodiment 1, glucose dehydrogenase-Ca 2+ Preparation of complex

[0036] 1. Prepare PBS buffer solution (pH=7.4) of glucose dehydrogenase with an enzyme concentration of 0.5 mg / mL and 100 mM calcium chloride aqueous solution.

[0037] 2. Take 2 mL of the enzyme solution in step 1 in a glass vial, add 0.02 mL of the calcium chloride solution in step 1, and mix well (in the mixed solution, Ca 2+ The mass ratio of glucose dehydrogenase to glucose dehydrogenase is 0.08:1) and then placed in an incubator at 4°C for 24 hours.

[0038] 3. Add 2wt% sucrose to the mixture obtained in step 2 as a lyoprotectant. The glucose dehydrogenase-calcium ion nanocomposite was obtained by freeze-drying for 12 hours.

[0039] The scanning electron micrograph of the nanocomposite prepared in this embodiment is as follows figure 1 shown by figure 1 It can be seen that the complex is a polyhedron of about 500 nm.

[0040] The enzyme loading (determined by BCA method to measure the remainin...

Embodiment 2

[0041] Embodiment 2, glucose oxidase-Zn 2+ Preparation of complex

[0042] The calcium chloride solution in the preparation step 1 of embodiment 1 is replaced with zinc chloride solution (50mM) respectively, and glucose dehydrogenase is replaced with glucose oxidase, Zn in the gained mixed solution 2+ The mass ratio to glucose oxidase is 0.065:1, no sucrose is added as a freeze-drying protective agent before freeze-drying, and other operations are the same.

[0043] Glucose Oxidase-Zn 2+ SEM images of nanocomposites / aggregates are shown in figure 2 shown by figure 2 It can be seen that the composite / aggregate presents a porous sheet-like structure.

[0044] Glucose oxidase-Zn prepared by the present embodiment 2+ The enzyme loading in the nanocomposite was 2.1%.

Embodiment 3

[0045] Embodiment 3, glucose dehydrogenase-Cu 2+ Preparation of complex

[0046] The calcium chloride solution in the preparation step 1 of embodiment 1 is replaced with copper sulfate solution (100mM), in the gained mixed solution Cu 2+ The mass ratio to glucose dehydrogenase is 0.128:1, no sucrose is added as a lyoprotectant before lyophilization, and other operations are the same.

[0047] Glucose dehydrogenase-Cu 2+ The scanning electron microscope image of the nanocomposite is shown in image 3 shown by image 3 It can be seen that the complex is a small ball of about 2um, which is formed by stacking polyhedrons.

[0048] Glucose dehydrogenase-Cu prepared by the present embodiment 2+ The enzyme loading in the nanocomposite was 2.3%.

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Abstract

The invention discloses an enzyme-metal ion nano compound and a preparation method thereof.The preparation method includes following steps: drying a water solution of metal ions and enzyme to obtain the enzyme-metal ion nano compound, wherein the metal ions are one of calcium ions, magnesium ions, iron ions, ferrous ions, copper ions, barium ions, manganous ions, bivalent nickel ions and zinc ions.An enzyme immobilization method has the advantages of simple operation and low cost.By controlling concentration of the metal ions and the enzyme in the solution, the enzyme-metal ion nano compound prepared by the method is not generated in a precipitate form while a nano compound of 500-1200 nm soluble in the water solution is formed, stability of the enzyme in the nano compound is improved remarkably, and enzyme activity is basically maintained unchanged and even is improved.Due to above characteristics, the enzyme-metal ion nano compound is different from conventional nano carrier immobilized enzyme, and has good application prospect in the field of industrial enzyme catalysis, biosensors and biomedicine.

Description

technical field [0001] The invention relates to an enzyme-metal ion nanocomposite and a preparation method thereof, belonging to the technical field of immobilized enzymes. Background technique [0002] Enzymes are highly efficient catalysts in nature, which are of great significance to the realization of environmentally friendly chemical production processes and efficient and sensitive analytical detection devices. However, due to the instability of their protein structures, natural enzymes will lose their activity due to conformational changes in the actual application environment, which limits their applications. The natural enzyme is immobilized on the carrier material. Based on the multi-point combination and restricted space interaction between the carrier material and the enzyme molecule, the protein structure can be stabilized, thereby improving the stability of the enzyme in practical applications. It also has good operability in application. Therefore, improving ...

Claims

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Application Information

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IPC IPC(8): C12N11/14
CPCC12N11/14C12N9/0006C12N9/0061C12N9/0065C12N9/0089C12Y101/03004C12Y101/05002C12Y110/03002C12Y111/01009C12Y115/01001
Inventor 戈钧吴晓玲王雪蕊
Owner TSINGHUA UNIV
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