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A new material for enriching and separating sialylated glycopeptides and its application

A technology of sialylated sugar and enrichment solution, which is applied in the field of enrichment and separation of sialylated glycopeptides, which can solve the problems of destroying the structure of sugar chains, difficult control of conditions, and many chemical reaction steps, etc., and achieves cheap and easy-to-obtain materials and specific enrichment High performance and remarkable effect

Active Publication Date: 2020-06-19
THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method has many chemical reaction steps, the conditions are not easy to control, and the chemical reaction process destroys the structure of the sugar chain, which is not conducive to further identification and analysis of the sugar chain type and structure.
However, zwitterionic hydrophilic chromatography and strong ion exchange chromatography can non-selectively enrich various types of glycopeptides, but the specificity is poor, the enrichment efficiency needs to be improved, and it is easy to cause the loss of sialic acid information

Method used

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  • A new material for enriching and separating sialylated glycopeptides and its application
  • A new material for enriching and separating sialylated glycopeptides and its application
  • A new material for enriching and separating sialylated glycopeptides and its application

Examples

Experimental program
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Effect test

Embodiment 1

[0030] Embodiment 1, application g-C 3 N 4 Enrichment of sialylated glycopeptides

[0031] 1. g-C 3 N 4 Procedure for Enrichment of Sialylated Glycopeptides

[0032] Step 1, g-C 3 N 4 material preparation

[0033] Grind the precursor (which can be dicyandiamide or melamine or urea) and add it to the crucible, put it in a muffle furnace at 550°C and cover it for calcination for 4 hours, take out the light yellow product, and grind it to get g-C 3 N 4 powder.

[0034] Step 2, preparation of enzymatic hydrolysis sample solution

[0035]Take 20 μL of 1 μg / μL protein (it can be bovine fetuin or transferrin or bovine fetuin, transferrin and bovine albumin mixed protein), add 2.2 μL 250 mM ammonium bicarbonate and 0.6 μL 1M dithiothreitol (25 mM bicarbonate ammonium), incubate at 56°C for 60 minutes (reduction reaction, the purpose is to open the disulfide bond), then add 5.6 μL containing 500mM iodoacetamide (prepared with 25mM ammonium bicarbonate), mix thoroughly, and in...

Embodiment 2

[0071] Example 2, g-C 3 N 4 Morphological and structural characterization of enriched materials

[0072] In this example, for g-C 3 N 4 Enriched materials for morphology and structural characterization, and further speculated that g-C 3 N 4 Mechanism of enrichment for sialylated glycopeptides.

[0073] Image 6 is g-C 3 N 4 The scanning electron microscope image can be found g-C 3 N 4 It is a layered micron-scale bulk phase structure, and a layered structure below 50nm can be seen in some positions. Figure 7 a and Figure 7 b is g-C respectively 3 N 4 TEM and high-resolution TEM images of . As shown, g-C 3 N 4 It is formed by the accumulation of amorphous single layers.

[0074] Figure 8 a Illustration is g-C 3 N 4 It can be seen that there are a large number of triazine rings and N-H bonds in the structure. Figure 8 a is g-C 3 N 4 The X-ray photoelectron spectrum of the X-ray photoelectron spectrum, the peaks at 286ev and 397ev correspond to the bind...

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Abstract

The invention discloses a method for specifically enriching sialylated glycopeptides, and relates to applications of g-C3N4 in enrichment of sialylated glycopeptides, and / or sialylated polysaccharides, and / or sialylated glycosides. The method for specifically enriching sialylated glycopeptides comprises: (1) adding a g-C3N4 dispersion liquid into the enzyme digestion product of sialylated glycoprotein, and carrying out vibration enrichment on sialylated glycopeptides; and (2) carrying out centrifugation on the enrichment liquid, and adding an eluent to the g-C3N4 precipitate so as to obtain the sialylated glycopeptide solution. According to the present invention, the method based on the g-C3N4 material with characteristics of environmental protection, simple preparation and low cost has advantages of mild enrichment condition and high specificity, can retain the structural information of the sialylated saccharide chain to the maximum extent, can truly reflect the natural glycosylationstatus of proteins, and has important application values in the detection of sialylated glycopeptides in complex samples and the screening of related tumor markers.

Description

technical field [0001] The invention relates to a new material and a new method for enriching and separating sialylated glycopeptides. Based on graphitic carbon nitride (g-C 3 N 4 ) The characteristic that a large number of N atoms in the unique triazine ring structure of the material can form hydrogen bonds with the sialylated sugar chain O-H, and the unique π interaction adsorption performance of the conjugated structure, realize the sialylated sugar without destroying the glycoprotein structure. Efficient, low-cost, specific enrichment separation of peptides, sialylated glycans, or sialylated glycosides. It provides new ideas and methods for the detection of sialylated glycopeptides in complex samples and the screening of related tumor markers. Background technique [0002] In the life activities of organisms, sialylated sugar chains play a role in specific recognition and mediation, and have an extremely important impact on the structure and function of proteins. Stu...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/34
Inventor 李智立张沫
Owner THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI
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