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A method for enriching sialylated glycopeptides

A sialylated sugar and sialylated technology, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of increasing the difficulty of sugar chain analysis and poor specificity, and achieve the effect of significant enrichment effect, high specificity and low cost

Active Publication Date: 2016-08-17
THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Strong cation chromatography and zwitterionic hydrophilic interaction liquid chromatography have been proven to be widely used for the enrichment and separation of various sialylated glycopeptides and / or sialylated glycans, but the cumbersome operation process will cause sialylated sugars The hydrolysis of peptides and / or sialylated glycans and the introduction of a large number of inorganic salt ions increase the difficulty of subsequent sugar chain analysis. In addition, the specificity of this method is poor

Method used

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  • A method for enriching sialylated glycopeptides
  • A method for enriching sialylated glycopeptides
  • A method for enriching sialylated glycopeptides

Examples

Experimental program
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Effect test

Embodiment 1

[0035] Example 1. Enrichment of sialylated glycopeptides using Populus tomentosa flocs

[0036] 1. Preparation of sample solution

[0037] 1. Preparation of sample solution A

[0038]Weigh 20 μg bovine fetuin, dissolve in 100 μL aqueous solution containing 25 mM ammonium bicarbonate and 50 mM dithiothreitol, incubate at 56 ° C for 45 minutes (reduction reaction, the purpose is to open the disulfide bond), then add 100 μL containing 200 mM iodine Aqueous solution of acetamide and 25mM ammonium bicarbonate, mix thoroughly, and react in the dark at 25°C (room temperature is sufficient) for 45 minutes (to block the disulfide bond opened in the reduction reaction), and then add 10μL of 10mM dithiothreitol aqueous solution to terminate After the reaction, 0.4 μg of trypsin was added and digested at 37°C for 12 hours, then the system was freeze-dried, and then the dry matter was dissolved in 30 μL of 80% acetonitrile aqueous solution to obtain sample solution A.

[0039] 2. Prepara...

Embodiment 2

[0080] Example 2. Structural characterization of poplar catkins and investigation of its active ingredients for enrichment of sialylated glycopeptides

[0081] In this example, poplar catkins, the filling medium of Popcat-tip, were structurally characterized, and the active ingredients enriched in sialylated glycopeptides in poplar catkins were further analyzed.

[0082] The scanning electron microscope analysis results of poplar catkins and absorbent cotton are as follows: image 3 shown. The diameter range of poplar catkins is 6-8 μm, which is obviously smaller than that of absorbent cotton (20-25 μm), indicating that poplar catkins have a relatively large specific surface area per unit volume, thereby increasing the unit contact area with the analyte, which is beneficial to Poplar catkins recognize and combine with the target. In addition, the surface of poplar floc is relatively smooth, while the surface of absorbent cotton is relatively rough. It can be inferred that th...

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Abstract

The present invention discloses a method for enriching sialylaglycopeptide, sialylated glycans or sialylated glycoside. The requirements of the invention protect application of Chinese white poplar catkin to enrichment of sialylaglycopeptide, sialylated glycans or sialylated glycoside. The invention also protects a method for enriching sialylaglycopeptide. The method comprises the steps of: (1) filling a column container with Chinese white poplar catkin to obtain a Popcat column; and (2) enriching sialylaglycopeptide from the digestion products of sialylated glycoprotein by using the Popcat column. The method provided by the invention has the advantages of mild conditions, low cost of column filling material, high singularity and substantial enrichment effect. The enrichment method provided by the invention can maintain the structure information of configuration information sialylated carbohydrate chain or glycan, preserve intact specific peptide information; the information is conducive to subsequent mass spectrometric analysis and reflects the native glycosylation state of protein and has great significance for the research on clinical diagnostic markers and disease mechanism.

Description

technical field [0001] The present invention relates to a method for enriching sialylated glycopeptides, sialylated glycans or sialylated glycosides. Background technique [0002] Sialic acid is an extremely important class of neuraminic acid derivatives. It is usually connected to the end of glycans, and it is one of the important material bases for the diversification of glycan structures and functions. It is widely involved in the targeted transmission of signaling molecules, such as ligands Targeted recognition and adhesion of receptors, antigens / antibodies, and enzymes / substrates, abnormal sialylated glycans can lead to the occurrence and development of diseases. Studies have shown that abnormal physiological states (such as abnormal brain development, cancer metastasis, viral infection, cardiac arrhythmia, etc.) are closely related to abnormal levels of sialylated glycans of some molecules in the body. Sialylated glycoproteins are a class of biomolecules that are comm...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N30/08G01N30/72
Inventor 李智立王晓东
Owner THE INST OF BASIC MEDICAL SCI OF CHINESE ACAD OF MEDICAL SCI
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