Protein assembly and preparing method thereof

A protein and assembly technology, applied in the field of protein assembly and its preparation, can solve problems such as inability to determine protein reaction sites, complex steps, etc., and achieve the effects of good salt ion and protein molecule interference and good stability.

Active Publication Date: 2018-10-12
TECHNICAL INST OF PHYSICS & CHEMISTRY - CHINESE ACAD OF SCI +1
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] In addition, there are many reactive sites on the surface of protein molecules, and common chemical modifications often cannot determine the reactive sites of proteins (such as Xin Huang, et al.Interfacial assembly of protein-polymer nano-conjugates into stimulus-responsive biomimetic protocells[J] .Nat.Commun.2013, 4, 2239.)
Modification of specific sites of proteins by chemical methods often requires complex steps

Method used

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  • Protein assembly and preparing method thereof

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preparation example Construction

[0049] In another aspect, the present invention also provides a method for preparing the above-mentioned protein assembly, comprising the following steps:

[0050] 1) uniformly disperse the virus nanoparticles in a neutral buffer solution, and stir to form a reaction solution 1;

[0051] 2) Add 1-hydroxybenzotriazole (HOBT) solution and fluorine tail solution to reaction solution 1, stir to form reaction solution 2;

[0052] 3) Adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) solution in portions to reaction solution 2, stirring to form reaction solution 3;

[0053] 4) The reaction solution 3 is dialyzed to obtain virus nanoparticles with surface-modified fluorine tails;

[0054] 5) adding the surface-modified fluorine-tailed virus nanoparticles into the acetic acid solution, centrifuging after standing, and taking the supernatant;

[0055] 6) The supernatant is desalted, the desalted solution is detected and the protein-containing part is collected; ...

Embodiment 1

[0076] Add 1 mL of HEPES buffer solution with an ionic strength of 1M and a pH value of 7.4, 5.2 mL of ultrapure water and 1.7 mL of DMSO into the bottle, and stir and mix evenly at 4°C. After cooling down, add 0.5 mL of tobacco mosaic virus aqueous solution (total 10 mg), add 193 μL of HOBT / DMSO solution (20 mg / mL) and 1.2 mL of NH 2 CH 2 CF 2 CF 2 CF 3 / DMSO (10mg / ml) solution, after stirring for 30 minutes, add 91μL of EDC aqueous solution (10mg / mL), stir at 4°C, add 91μL of EDC aqueous solution after 12 hours and 20 hours respectively, and the reaction is completed in 24 hours. The dialysis bag is dialyzed in ultrapure water, and the water is replaced 5-6 times to obtain the tobacco mosaic virus with a fluorine tail (containing seven fluorine atoms) modified on the inner surface.

[0077] Add 1.6mL of glacial acetic acid to the centrifuge tube, add 0.8mL of fluorine-tailed tobacco mosaic virus on the inner surface to the glacial acetic acid, place it in a refrigerator ...

Embodiment 2

[0081] Add 1 mL of HEPES buffer solution with an ionic strength of 1 M and a pH value of 7.4, 5.2 mL of ultrapure water and 1.7 mL of DMSO into the bottle, and stir and mix evenly at 4°C. After cooling down, add 0.5 mL TMV aqueous solution (total 10 mg), add 193 μL HOBT / DMSO solution (20 mg / mL) and 0.8 mL NH 2 CH 2 CF 2 CF 3 / DMSO (10mg / ml) solution, stirred for 30 minutes, added 91 μL EDC aqueous solution (10mg / mL), stirred at 4°C, added 91 μL EDC aqueous solution after 12 hours and 20 hours, respectively. After 24 hours of reaction, the dialysis bag with a molecular weight cut-off of 1000 kDa was dialyzed in ultrapure water, and the water was replaced 5-6 times to obtain the tobacco mosaic virus with a modified fluorine tail (containing five fluorine atoms) on the inner surface.

[0082] Add 1.6mL of glacial acetic acid to the centrifuge tube, add 0.8mL of fluorine-tailed tobacco mosaic virus on the inner surface to the glacial acetic acid, place it in a refrigerator at 4°...

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Abstract

The invention discloses a protein assembly and a preparing method thereof. The protein assembly is formed by self-assemblying fluorine tail-modified virus nanoparticle capsid protein and is of a vesicle structure. According to the preparing method of the protein assembly, self-assembling characteristics of the virus nanoparticles are used, modification of special sites of the protein is achieved through a simple and effective method, and thus the protein assembly of the capsid protein modified with the fluorine tails with different lengths is successfully prepared. According to the protein assembly, the fluorine tails are selected to replace polymers and be used as chain segments for driving the protein molecules to be self-assembled, and the fluorine tails not only have mono-disperse molecular weight and clear molecular structure, but also form a hydrophobic and oleophobic third phase-fluorine phase through fluorine- fluorine interaction. Introduction of the fluorine tails cannot cause damage to monodispersion of the protein molecules, and can make the protein assembly have better stability in blood circulation.

Description

technical field [0001] The invention relates to the field of biomedical materials. More specifically, it relates to a protein assembly and its preparation method. Background technique [0002] Definite molecular weight and clear molecular structure are important factors for the application transformation of biomedical materials. The protein assemblies inspired by nature not only have definite molecular weight and clear molecular structure, but also have good biocompatibility. Protein assemblies are of great significance in the field of biomedicine, such as biosensing, drug delivery, and simulation of primitive cell functions (such as X.-H.Dong, et al.Three‐Dimensional Ordered Antibody Arrays Through Self‐Assembly of Antibody -Polymer Conjugates [J]. Angew. Chem. Int. Ed. 2017, 56, 1273. Ge, J., et al. Protein - Polymer Hybrid Nanoparticles for Drug Delivery [J]. Small, 2012, 8, 3573.). However, proteins such as bovine serum albumin generally do not have the ability to sel...

Claims

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Application Information

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IPC IPC(8): C07K14/08
CPCC07K14/005C12N2770/00023C12N2770/00051
Inventor 牛忠伟高偲嘉田野
Owner TECHNICAL INST OF PHYSICS & CHEMISTRY - CHINESE ACAD OF SCI
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