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Magnetic nano-material, preparation method and purpose thereof

A technology of magnetic nano and nano materials, applied in the field of biochemical analysis, can solve problems such as inability to analyze and false positives

Inactive Publication Date: 2017-03-01
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The research in the prior art basically uses biotin or metabolic labeling methods to enrich such modifications. However, practice shows that using biotin methods is easy to obtain endogenous biotinylated proteins / peptides, resulting in false positives As a result, only living cells can be analyzed using metabolic markers, and samples such as tissue samples and body fluids cannot be analyzed

Method used

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  • Magnetic nano-material, preparation method and purpose thereof
  • Magnetic nano-material, preparation method and purpose thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0051] Preparation of Novel Magnetic Nanomaterials Encapsulated by N-Hydroxysuccinimide 3-(2-pyridyldimercapto)propionate

[0052] 1. Synthesis of Amino Magnetic Beads

[0053] 1) Weigh 8 g of anhydrous sodium acetate (NaAc) and iron trichloride (FeCl 3 ·6H 2 (0) 2g in a beaker, add 60ml ethylene glycol, stir and dissolve;

[0054] 2) Ethylenediamine is a waxy solid, which can be dissolved by heating and becomes liquid, and 7.2ml (about 7.2g) is sucked into the above-mentioned reaction beaker with a straw. After the dissolution is complete, the solution turns red-orange;

[0055] 3) Pour the dissolved solution into the reactor;

[0056] 4) Put the reactor into an oven, and react at 180°C for 6 hours;

[0057] 5) After the reaction is over, take it out and cool it;

[0058] 6) washing the material with water and ethanol respectively;

[0059] 2. Synthesis of Fe 3 o 4 @NH 2 @SPDP

[0060] 1) Take the amino magnetic beads Fe synthesized above 3 o 4 @NH 2 , wash with...

Embodiment 2

[0068] 1) Take 1 μl of peptide S-G#R (100ng / μl) with palmitoylation modification and Fe 3 o 4 @NH 2 @SPDP was incubated in a PBS-EDTA solution of hydroxylamine (pH 7.4). After 4 hours at room temperature, the supernatant was removed by magnetic separation. After the material was washed, the bound S- GR, use C18ziptip to desalt the eluent. At this time, the S-GR peptide has lost the palmitoyl-modified side chain, and the molecular weight has changed. After the target is analyzed by MALDI-TOF MS, the results are as follows Figure 5 as shown, Figure 5 (a) and Figure 5 (b) show respectively with Fe 3 o 4 @NH 2 The mass spectrograms before and after @SPDP enrichment, after enrichment, a clear 994 peptide peak can be obtained;

[0069] 2) After mixing the S-G#R peptide and the MYO enzymatic peptide according to the molar ratios of 1:1, 1:20, 1:50 and 1:100, respectively, 1 μl of the mixture was used for MALDI-TOF MS analysis. Such as Figure 6 (a)(b) and Figure 7 (a) A...

Embodiment 3

[0072] Take 1μl S-G#R (0.05ng / μl) and Fe 3 o 4 @NH 2 After incubation with @SPDP, evaluate the enrichment sensitivity of the method for palmitoylated peptides, and sample the target from the enriched and eluted solution for MALDI-TOFMS analysis (such as Figure 9 Shown), the S-GR signal peak is obvious after enrichment, indicating that the method has high sensitivity.

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Abstract

The invention belongs to the field of biochemical analysis, and relates to a magnetic nano-material, a preparation method and a purpose of the magnetic nano-material for a method for enrichment palmitoyl-modified protein / peptide segment. The method employs a synthesized nano Fe3O4 magnetic core as a material magnetic center, 3-(2-pyridine dimercapto)propionic acid N-hydroxysuccinimide eater (SPDP) is used for coating the Fe3O4 magnetic core, and a Fe3O4-coated NH2-coated SPDP magnetic nano-material is prepared: the nano-material can selectively increase the efficiency of enrichment of palmitoyl-modified protein / peptide segment, and can be used for analyzing a practical biological sample. The magnetic nano-material has the advantages of good selectivity, high sensitivity, simple preparation, and rapid and convenient separation, and the enrichment method of the magnetic nano-material has the advantages of simple operation, time saving and economy, and high efficiency and sensitivity.

Description

technical field [0001] The invention belongs to the field of biochemical analysis, and relates to magnetic nanomaterials, in particular to a magnetic nanomaterial, a preparation method thereof, and an application in a method for enriching palmitoylated modified proteins / peptides. [0002] technical background [0003] The prior art discloses that S-acylation modification of proteins is a very important form of post-translational modification. S-acylation (S-acylation) modification of protein, also known as palmitoylation modification (palmitoylation), it refers to the covalent modification of long-chain fatty acids (mainly 16-carbon palmitic acid) to on the sulfhydryl group of protein cysteine. Studies have shown that S-acylation modification is dynamic and reversible, it can regulate protein activity and stability, protein-protein interaction, effectively increase protein hydrophobicity, promote self or other proteins to localize membrane lipid rafts, etc., in It plays an ...

Claims

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Application Information

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IPC IPC(8): G01N1/40H01F1/42
Inventor 陆豪杰张晓勤方彩云
Owner FUDAN UNIV
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