Laccase modification method on basis of amino acid terminal carboxyl group and application thereof

A technology of terminal carboxyl group and amino acid, applied in the application field of modified laccase in the modification of regenerated plant fiber, to achieve the effect of improving laccase activity, reducing the amount of laccase used, and simple operation steps

Inactive Publication Date: 2013-12-11
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] One of the purposes of the present invention solves the existing problems and deficiencies in the existing laccase modification, and proposes a new modification method for improving laccase activity and stability

Method used

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  • Laccase modification method on basis of amino acid terminal carboxyl group and application thereof
  • Laccase modification method on basis of amino acid terminal carboxyl group and application thereof
  • Laccase modification method on basis of amino acid terminal carboxyl group and application thereof

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Effect test

Embodiment 1

[0025] Modification step: Mix 5mL, 10mg / mL laccase solution (laccase SUKALacc produced by Aspergillus) with 0.4mg / mL, 5mL L-phenylalanine methyl ester hydrochloride solution, add 1.0 mg1-Ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride 1-Ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC), reacted at 25°C for 4h under magnetic stirring conditions, Wherein the laccase solution and the L-phenylalanine methyl ester hydrochloride solution all use the potassium dihydrogen phosphate / dipotassium hydrogen phosphate (KH 2 PO 4 / K 2 HPO 4 ) buffer solution preparation. After the reaction is completed, transfer the mixed solution into a dialysis bag, and dialyze for 48 hours at room temperature in a dipotassium hydrogen phosphate / citric acid buffer solution with a pH of 5.0 and a concentration of 0.2 mol / L to remove excess L-phenylalanine methyl ester hydrochloride and EDC, to obtain a modified laccase solution, dilute to 25mL, freeze and store for later use, and ...

Embodiment 2

[0041]Mix 5 mL, 10 mg / mL laccase solution (laccase SUKALacc produced by Aspergillus) with 0.2 mg / mL, 6 mL L-phenylalanine methyl ester hydrochloride solution, and add 0.8 mg 1-ethyl-( 3-Dimethylaminopropyl) carbodiimide hydrochloride 1-Ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC), reacted at 25°C for 4h under magnetic stirring conditions, in which the laccase solution Both potassium dihydrogen phosphate / dipotassium hydrogen phosphate (KH 2 PO 4 / K 2 HPO 4 ) buffer solution preparation. After the reaction is completed, transfer the mixed solution into a dialysis bag, and dialyze at room temperature for 24 hours in a dipotassium hydrogen phosphate / citric acid buffer solution with a pH of 5.0 and a concentration of 0.2 mol / L to remove excess L-phenylalanine methyl Ester hydrochloride and EDC to obtain a modified laccase solution, dilute to 25mL, and freeze for future use. The laccase activity and half-life were determined according to the method described ...

Embodiment 3

[0044] Mix 5 mL, 10 mg / mL laccase solution (laccase SUKALacc produced by Aspergillus) with 0.1 mg / mL, 10 mL L-phenylalanine methyl ester hydrochloride solution, and add 1.0 mg 1-ethyl-( 3-Dimethylaminopropyl) carbodiimide hydrochloride 1-Ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC), reacted at 25°C for 4h under magnetic stirring conditions, in which laccase solution and L - Phenylalanine methyl ester hydrochloric acid solution is all used potassium dihydrogen phosphate / dipotassium hydrogen phosphate (KH 2 PO 4 / K 2 HPO 4 ) buffer solution preparation. After the reaction is completed, transfer the mixed solution into a dialysis bag, and dialyze at room temperature for 12 hours in a dipotassium hydrogen phosphate / citric acid buffer solution with a pH of 5.0 and a concentration of 0.2 mol / L to remove excess L-tryptophan methyl ester hydrochloride and EDC to obtain a modified laccase solution, which was adjusted to 25 mL, and stored frozen for later use. Th...

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Abstract

The invention provides a modification method on the basis of crosslinking reaction between laccase amino acid residue terminal carboxyl group (-COOH) and amino group of methyl L-phenylalaninate hydrochloride to enhance activity and stability of the laccase, which comprises the following step: mixing laccase, methyl L-phenylalaninate hydrochloride and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride at a certain temperature to react to obtain the modified laccase. Compared with the unmodified laccase, the modified laccase has higher activity and stability. The modified laccase can be used for aftergrowth fiber modification, and can save the laccase consumption on the premise of enhancing the strength property of the paper sheets made from aftergrowth fibers.

Description

technical field [0001] The invention relates to the fields of pulping, papermaking and bioengineering, in particular to a modification method for improving the activity and stability of laccase and the application of the modified laccase in the modification of regenerated plant fibers. Background technique [0002] Compared with native plant fibers, the swelling performance, fiber strength and paper physical properties of regenerated plant fibers are all decreased. The reduction of the strength of regenerated plant fibers has greatly affected its recycling. In order to improve the strength properties of regenerated plant fibers, improve their utilization level and value, it is necessary to modify them. Among the many methods for modifying regenerated plant fibers, laccase and laccase mediator system is currently an ideal biomodification method for regenerated plant fibers. Its advantages include: (1) mild reaction conditions; (2) little damage to fibers; (3) no secondary po...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/02D21H17/22
Inventor 万金泉陈杨梅张全升马邕文王艳黄明智
Owner SOUTH CHINA UNIV OF TECH
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