Method for enriching and purifying glycosylation peptides by nano chitosan derivative

A technology of nano-chitosan and derivatives, applied in the field of biomedical nano-materials, can solve the problem of no enrichment of glycopeptides, and achieve the effects of good biocompatibility, cheap and easy-to-obtain materials, and large specific surface area.

Inactive Publication Date: 2015-04-22
PEKING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] In the patent ZL 200910086838.6, a nano-chitosan derivative containing phenylboronic acid, a preparation method and its application in enrichment and purification of glycosylated polypeptide / protein are reported; Glycopeptides at the Fc end have no enrichment effect

Method used

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  • Method for enriching and purifying glycosylation peptides by nano chitosan derivative
  • Method for enriching and purifying glycosylation peptides by nano chitosan derivative
  • Method for enriching and purifying glycosylation peptides by nano chitosan derivative

Examples

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Effect test

Embodiment 1

[0028] Enrichment and Analysis of Horseradish Peroxidase (HRP) Glycosylated Peptides

[0029] 1) Preparation of sample solution: Add 25 μg horseradish peroxidase (Sigma) to 10 μL containing 8M urea, ethylenediaminetetraacetic acid (EDTA) and 10 mM tris(2-chloroethyl)phosphate (TCEP), shake at room temperature After 1 hour, add 40 μL of 50 mM ammonium bicarbonate solution (pH 8.2), add trypsin according to the mass ratio of trypsin (40:1) for enzymolysis reaction, and control the enzymolysis temperature at 37°C. After overnight reaction, 2% trifluoroacetic acid (TFA) was added to terminate the reaction. The obtained proteolysis solution was stored in a -80°C refrigerator for future use.

[0030] 2) Enrichment of glycosylated peptides and MALDI-TOF mass spectrometry analysis: 2 μL 1×10 -6 The mM horseradish peroxidase enzymolysis solution was dissolved in 100 μL sample solution, wherein the sample solution was an aqueous solution containing 80% acetonitrile and 5% trifluoroace...

Embodiment 2

[0033] The same method as in Example 1, wherein step 2) eluting the enriched glycosylated polypeptide with 5% to 10% ammonia solution or an aqueous solution containing 10% to 50% acetonitrile and 5% formic acid, concentrated and dried, Add 2 μL of 0.1% trifluoroacetic acid aqueous solution to dissolve, pipette 0.5 μL containing 1% HO 3 PO 4 50% acetonitrile DHB (25mg / mL) matrix solution was spotted on the target plate, 0.5 μL of the above solution containing glycosylated polypeptide was added and mixed, and then MALDI-TOF mass spectrometry analysis was performed.

Embodiment 3

[0035] The method is the same as in Example 2, wherein the mass spectrometer used is nano-LC-ESI-MS, and is carried out according to a conventional method in the art.

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Abstract

The invention discloses a method for enriching and purifying glycosylation peptides by a nano chitosan derivative, which belongs to the technical field of biomedical nano-material. The particle size of the nano chitosan derivative is 1-300nm, a solid supported matrix is chitosan, the derivative has an active carboxyl function group and enables complexation with transition metal Ti <4+> ion. The nano chitosan derivative contains a sugar chain and metal Ti <4+> ion, so that glycosylation polypeptides can be enriched and purified with high selectivity and specificity. In addition, the nano chitosan derivative can enrich the glycosylation polypeptides (such as IgG) containing sialic acid as well as enrich polypeptides (such as HRP) of different glycosyls. The nano chitosan derivative can be used for enriching and purifying glycosylation polypeptides with low abundance in a biological sample, and can be used for biological and medical fields comprising clinical diagnosis.

Description

technical field [0001] The invention belongs to the technical field of biomedical nanomaterials, and in particular relates to a method for enriching and purifying glycosylated polypeptides with nano-chitosan derivatives. Background technique [0002] Post-translational protein modification is a hot topic in proteomics research, and protein glycosylation is one of the most common and important ways of protein post-translational modification. Glycosylation of protein almost regulates the whole process of life activities, including cell proliferation, development and differentiation, metabolism, immune response, tumorigenesis and so on. Some glycosylated proteins are used as relevant targets or biomarkers for the treatment of diseases for the early detection and identification of cancer, such as carcinoembryonic antigen for the detection of rectal cancer, breast cancer, prostate cancer and lung cancer; CA-125 for the detection of ovarian cancer ; Specific prostate carcinoembry...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/28G01N1/34G01N30/08G01N30/06
Inventor 邹霞娟杨彬
Owner PEKING UNIV
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