Method for assisting lysozyme renaturation in vitro by granular poly(N-isopropylacrylicamide-sodium acrylate) copolymer hydrogel

A technology of isopropyl acrylamide and sodium acrylate, applied in biochemical equipment and methods, enzymes, enzymes, etc., can solve the problems of weak mechanical strength, narrow adjustable range of phase transition temperature, and application limitations.

Inactive Publication Date: 2009-02-11
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the mechanical strength of the granular gel is weak, and the adjustable range of phase transition temperature is narrow, which limits its application.

Method used

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  • Method for assisting lysozyme renaturation in vitro by granular poly(N-isopropylacrylicamide-sodium acrylate) copolymer hydrogel
  • Method for assisting lysozyme renaturation in vitro by granular poly(N-isopropylacrylicamide-sodium acrylate) copolymer hydrogel
  • Method for assisting lysozyme renaturation in vitro by granular poly(N-isopropylacrylicamide-sodium acrylate) copolymer hydrogel

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] 1) Take 40 mL of NaOH solution with a concentration of 0.2 mol / L, add excess acrylic acid aqueous solution, stir and react for 4 hours, evaporate under reduced pressure to remove the solvent and residual acrylic acid, and collect the sodium acrylate solid;

[0021] 2) Dissolve the surfactant Tween-80 in 75 mL of organic phase liquid paraffin to make the concentration 1% (g / mL), add it into the reactor, feed nitrogen gas, and stir for 20 minutes to fully disperse the surfactant. Then the raw material aqueous solution that the volume is organic phase volume 1 / 3 containing various reaction components is slowly added dropwise in the reactor with pipette gun, and raw material aqueous solution is composed of main monomer concentration 14% (g / g), cross-linking degree is 10% (g / g), the molar ratio of monomer N-isopropylacrylamide to comonomer sodium acrylate is 15.7:1, the concentration of oxidant ammonium persulfate is 0.5% (g / mL), under nitrogen protection After stirring for ...

Embodiment 2

[0024] 1) Take 50 mL of NaOH solution with a concentration of 0.2 mol / L, add excess acrylic acid aqueous solution, stir and react for 4 hours, evaporate under reduced pressure to remove the solvent and residual acrylic acid, and collect the sodium acrylate solid;

[0025] 2) Dissolve the surfactant Tween-80 in 75 mL of organic phase liquid paraffin to make the concentration 1% (g / mL), add it into the reactor, feed nitrogen gas, and stir for 30 min to fully disperse the surfactant. Then the raw material aqueous solution that the volume is organic phase volume 1 / 3 containing various reaction components is slowly added dropwise in the reactor with pipette gun, and raw material aqueous solution is composed of main monomer concentration 14% (g / g), cross-linking degree 10% (g / g), the molar ratio of monomer N-isopropylacrylamide to self-made co-monomer sodium acrylate is 19:1, the concentration of oxidant ammonium persulfate is 0.5% (g / mL), in nitrogen Under protection, after stirrin...

Embodiment 3

[0028] 1) Take 60 mL of NaOH solution with a concentration of 0.4 mol / L, add excess acrylic acid aqueous solution, stir and react for 6 hours, evaporate under reduced pressure to remove the solvent and residual acrylic acid, and collect sodium acrylate solid;

[0029]2) Dissolve the surfactant Tween-80 in 100 mL of organic phase liquid paraffin to make the concentration 1.5% (g / mL), add it into the reactor, blow in nitrogen, and stir for 30 min to fully disperse the surfactant. Then the raw material aqueous solution that the volume is organic phase volume 1 / 3 containing various reaction components is slowly added dropwise in the reactor with pipette gun, and raw material aqueous solution is composed of main monomer concentration 14% (g / g), cross-linking degree is 10% (g / g), the molar ratio of monomer N-isopropylacrylamide to self-made co-monomer sodium acrylate is 9:1, and the concentration of oxidant ammonium persulfate is 0.5% (g / mL). Under protection, after stirring for 40 ...

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Abstract

The invention discloses a method for assisting the in vitro renaturation of lysozyme by using granular poly (N-isopropylacrylamide-sodium acrylate) copolymer gel, the specific steps are as follows: 1) the granular poly (N-isopropylacrylamide-sodium acrylate) copolymer gel is prepared by using the inverse suspension copolymerization; 2) the granular poly (N-isopropylacrylamide-sodium acrylate) copolymer gel is used for assisting the in vitro renaturation of the lysozyme. The granular poly (N-isopropylacrylamide-sodium acrylate) copolymer gel which is developed by the method is taken as a novel protein in vitro renaturation additive, thereby having the following advantages: (1) the activity recovery rate of a target protein can be significantly improved under the condition of high denatured protein concentration; (2) the separation and the recovery are convenient by utilizing the thermosensitive characteristic of the gel after the completion of the renaturation, and the gel can be repeatedly utilized; (3) the hydrophobic performance of the gel can be regulated by controlling the content of the co-monomer sodium acrylate, thereby changing the low critical dissolution temperature and the largest swelling ratio and being applicable to the different target proteins. Therefore, the gel has better application prospect in the field of the protein renaturation.

Description

technical field [0001] The invention relates to a method for assisting lysozyme in vitro renaturation by granular poly(N-isopropylacrylamide-sodium acrylate) copolymer gel. Background technique [0002] Genetic engineering technology is the main body of modern biotechnology, and its rapid development makes the heterogeneous expression of proteins an important way to develop the biomedical industry and obtain special chemicals. However, in the process of industrialization of genetically engineered proteins, such a problem is often faced: the recombinant expression product usually exists in the form of inclusion bodies without biological activity. How to efficiently refold the inclusion body into a protein with natural conformation and biological activity is the key to determining whether the genetically engineered protein can be industrialized and utilized by humans. Therefore, the field of protein renaturation technology in vitro has become one of the research hotspots in re...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/36
Inventor 关怡新葛翔金佳钰姚善泾
Owner ZHEJIANG UNIV
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