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Method for coupling negative-charge protein with 3-aminopropyl triethoxy silane

An aminopropyltriethoxysilane and protein technology, which is applied in the field of protein modification or compound preparation of organic materials and biopolymer chemistry, can solve problems such as protein self-coupling, and achieve high protein modification efficiency and high stability , Modified uniform effect

Active Publication Date: 2019-09-10
SHANGHAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At the same time, the protein itself has amino and carboxyl groups, if the pH of the system is not well controlled, it may lead to self-coupling of the protein

Method used

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  • Method for coupling negative-charge protein with 3-aminopropyl triethoxy silane
  • Method for coupling negative-charge protein with 3-aminopropyl triethoxy silane
  • Method for coupling negative-charge protein with 3-aminopropyl triethoxy silane

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Effect test

Embodiment 1

[0030] In this embodiment, a method for coupling 3-aminopropyltriethoxysilane to a negatively charged protein comprises the following steps:

[0031] a. Activation stage:

[0032] At room temperature, in a 50ml eggplant-shaped bottle, 15mg of BSA was dissolved in 2ml of PB, wherein the pH of PB=6.5, and the concentration of PB was 0.1M to obtain a BSA buffer mixture, and then the BSA buffer mixture was transferred to In the chromatographic cabinet at 4°C, control the speed at 750rpm to mix the BSA buffer mixture evenly; then add 4.4mg of EDC and 6.6mg of NHS to the BSA buffer mixture in a 50ml eggplant bottle, and stir magnetically for 30min. To obtain a mixed liquid system, maintain the pH of the mixed liquid system between 4.5 and 7.2, and at the same time make the difference between the pH of the mixed liquid system and the isoelectric point of the protein more than 1.0, so that the protein is evenly dispersed in the mixed liquid system, and the protein Coagulation and sel...

Embodiment 2

[0042] This embodiment is basically the same as Embodiment 1, especially in that:

[0043] In this embodiment, a method for coupling 3-aminopropyltriethoxysilane to a negatively charged protein comprises the following steps:

[0044] a. Activation stage:

[0045] At room temperature, weigh 5 mg of Insulin and dissolve it in 1.8 ml of Tris-HCl buffer, wherein the pH of Tris-HCl buffer is 7.4, the concentration of Tris-HCl buffer is 0.1M, and then add 200 μl of 0.12 M hydrochloric acid. Then transfer the mixed solution of 2ml of dissolved Insulin to a 50ml eggplant-shaped bottle. At this time, adjust the rotating speed of the magnetic stirrer to 700rpm to mix the mixed solution of Insulin evenly; then add 6ml of Tris with a pH of 7.4 and a concentration of 0.1M -HCl buffer to adjust pH. Add 4.4mg of EDC and 6.6mg of NHS to the mixed solution of Insulin in the 50ml eggplant-shaped bottle, and magnetically stir for 30min to obtain a mixed solution system, maintain the pH of the...

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Abstract

The invention discloses a method for coupling negative-charge protein with 3-aminopropyl triethoxy silane and belongs to the technical field of inorganic materials and biology high-polymer chemistry.According to the method, by controlling the pH of the protein in the activation process and the process of coupling the protein with the 3-aminopropyl triethoxy silane, the pH in the activation process needs to be controlled within 4.5-7.2, the pH in the coupling process is 7-8, and it is also guaranteed that the difference of the pH of the solution and the isoelectric point of the protein is above 1. According to the method, the problems of protein coagulation and self-coupling in the protein modification process through APTS are solved. The method is high in modification efficiency and evenin modification, the stability of the protein under the non-physiological condition is obviously improved, and the basis is laid for preparation of a nano silicon dioxide encapsulation protein nano material.

Description

technical field [0001] The invention relates to a method for improving protein stability, in particular to a protein amidation modification method for preparing nano-silica-encapsulated proteins, which is applied to the technical field of protein modification or chemical compound preparation of organic materials and biopolymers. Background technique [0002] Protein is the material basis of life, without protein there is no life. In recent years, due to the diversity of protein functions, more and more protein products have been produced and applied to various fields. However, the inherent instability of natural proteins under non-physiological conditions, such as denaturation, enzymatic hydrolysis, etc., limits their application in industry or normal laboratory conditions. Therefore, the improvement of protein stability is imminent. [0003] Nano-silica encapsulation of proteins has been proven to be an effective method to improve protein stability. Catalyzed 3-aminoprop...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/107
CPCC07K1/1077
Inventor 曹傲能华杰王海芳
Owner SHANGHAI UNIV
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