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Method for improving stability of bacillus subtilis Zj016 aminopeptidase by chemical modification of succinic anhydride

A chemical technology of Bacillus subtilis and succinic anhydride, applied in the fields of food additives and enzyme preparations, can solve the problems of easy inactivation, short half-life, affecting the use effect of enzymes, etc., and achieve the effect of improving the properties of the enzyme and improving the thermal stability.

Inactive Publication Date: 2013-07-17
JIANGNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] As a kind of biocatalyst, enzymes seriously affect the use effect of enzymes due to their shortcomings such as short half-life and easy inactivation.

Method used

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  • Method for improving stability of bacillus subtilis Zj016 aminopeptidase by chemical modification of succinic anhydride
  • Method for improving stability of bacillus subtilis Zj016 aminopeptidase by chemical modification of succinic anhydride
  • Method for improving stability of bacillus subtilis Zj016 aminopeptidase by chemical modification of succinic anhydride

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Embodiment 1

[0044] Weigh 20 mg of aminopeptidase powder and dissolve it in 15 mL, 0.05 mol / L, pH 8.5 Tris-HCl buffer solution, then slowly add 10 mg of succinic anhydride (SA) to the mixture, stir to dissolve, During the process, 2mol / L NaOH solution was used to maintain the pH value of the reaction system at 8.0, and the reaction was carried out at 4°C for 1 hour. After the reaction, dialyzed overnight at 4°C with 0.05mol / L, pH 8.5 Tris-HCl buffer solution. To remove excess modifiers, obtain succinic anhydride-modified aminopeptidase (modified enzyme SA-AE); weigh 20 mg of aminopeptidase powder and dissolve in 15 mL, 0.05 mol / L, pH 8.5 Tris-HCl buffer Dialyze with 0.05 mol / L, pH 8.5 Tris-HCl buffer at 4°C overnight to obtain the original enzyme solution (AE), and store in a 4°C refrigerator. The enzyme activity retention rate of the modified SA-AE was 105.81%. Compared with the original enzyme solution, the optimum reaction temperature of the modified enzyme was increased from 60°C to 65...

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Abstract

The invention discloses a method for improving stability of bacillus subtilis Zj016 aminopeptidase by chemical modification of succinic anhydride, belonging to the technical field of enzyme preparation and food additive. The method is based on residual enzymatic activity and relative enzyme activity; the residual enzymatic activity of modification enzyme liquid is 105.81%; the average amino modification rate of succinic anhydride (SA) for aminopeptidase is 55.86%. Compared with the proenzyme liquid, the optimal reaction temperature of the modification enzyme liquid is improved to 65 DEG C from 60 DEG C; the optimal pH is improved to 9.5-10.0 from 8.5; more than 70% of activity can be maintained after maintaining the temperature for 5 hours at 70 DEG C, so that it is shown that the thermalstability is improved drastically; the kinetic parameter Km is reduced to 0.75mmol.L<-1>, and the affinity of the modified aminopeptidase for primer is improved notably. SA chemical modification is an effective method for notably improving affinity of the bacillus subtilis aminopeptidase for the primer and thermal stability of the bacillus subtilis aminopeptidase, improves application characteristic of the bacillus subtilis aminopeptidase and plays an active role in promoting industrial production and further application of the aminopeptidase.

Description

technical field [0001] The invention discloses a method for chemically modifying succinic anhydride to improve the stability of Bacillus subtilis Zj016 aminopeptidase, which belongs to the technical field of enzyme preparations and food additives. Background technique [0002] Aminopeptidase is a general term for a class of exopeptidases that cleave amino acid residues from the N-terminus of proteins or peptide chains. It can be used as an enzyme for compounding in the preparation of proteolysis products, which can be used for debittering, deep hydrolysis, and preparation of multifunctional active peptides. It can also be used as an N-terminal sequencing tool enzyme and an enzyme for medical diagnosis. [0003] As a kind of biocatalyst, enzymes seriously affect the use effect of enzymes due to their shortcomings such as short half-life and easy inactivation. Enzyme stability refers to the ability of enzyme molecules to resist the influence of various factors, maintain a cer...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/96
Inventor 田亚平张静
Owner JIANGNAN UNIV
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