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Phosphorylated peptide adsorption enrichment method, additive and application

A technology for phosphorylated peptides and additives, applied in the field of protein and peptide enrichment and purification, can solve problems such as poor effect, affecting the accuracy and stability of phosphorylated peptides, and achieve increased adsorption selectivity, accuracy and stability. , reduce the effect of adsorption

Active Publication Date: 2014-10-29
SHENZHEN HUADA GENE INST
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  • Summary
  • Abstract
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AI Technical Summary

Problems solved by technology

However, the effect of existing non-specific inhibitors in phosphorylated peptide enrichment is not good, especially in the currently commonly used titanium dioxide enrichment method, the enrichment selectivity of existing non-specific inhibitors is generally 60% -70% or so; that is, there are still 30%-40% or more non-phosphorylated peptides enriched, which seriously affects the accuracy and stability of phosphorylated peptide detection

Method used

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  • Phosphorylated peptide adsorption enrichment method, additive and application
  • Phosphorylated peptide adsorption enrichment method, additive and application
  • Phosphorylated peptide adsorption enrichment method, additive and application

Examples

Experimental program
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Embodiment

[0040] In this example, mouse liver samples were taken as the research object, and phosphorylated peptides were enriched by the titanium dioxide enrichment method. Specific enrichment methods include:

[0041] First, glycolic acid, a routinely used non-specific adsorption inhibitor, was added to the loading buffer. At the same time, part of the sample to be tested extracted from the mouse liver was dissolved in the loading buffer, and the phosphorylated Enrich the peptide; collect the enriched product, analyze it, calculate the abundance of 20 amino acids in the non-phosphorylated peptide as the first abundance, and calculate the abundance of the 20 amino acids in the protein where the non-phosphorylated peptide is located as the For the second abundance, calculate the ratio of the first abundance to the second abundance, and select at least one amino acid with a higher ratio as an additive; then add the obtained additive to the loading buffer, and use the loading buffer with ...

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Abstract

The invention discloses a phosphorylated peptide adsorption enrichment method comprising (1) a non specific adsorption inhibitor and part of a to-be-tested sample are added a sample loading buffer, and an enrichment filter column is used for phosphorylated adsorption enrichment; (2) enrichment products are collected, the abundance of 20 kinds of amino acids in non phosphorylated peptide can be calculated as first abundance, the abundance of the non phosphorylated peptide in the 20 kinds of amino acids can be calculated as second abundance, at least one amino acid with higher ratio of first abundance to second abundance can be selected as an additive; (3) the sample loading buffer added with the additive is used for phosphorylated adsorption enrichment of the to-be-tested sample. By use of the competitive adsorption effects of amino acids and peptides, adsorption of the non phosphorylated peptide can be reduced, adsorption selectivity of the phosphorylated peptide enrichment can be increased, and the method lays the foundation for improving the accuracy and stability of phosphorylation detection.

Description

technical field [0001] This application relates to the field of enrichment and purification of proteins and polypeptides, in particular to an adsorption enrichment method for phosphorylated peptides, an additive for phosphorylation enrichment, a preparation method and application of the additive. Background technique [0002] In biological cells, proteins are the main function performers. Due to functional needs, many proteins will covalently bind various non-peptide groups on the amino acid chain during or after translation to form post-translational modifications. Phosphorylation of proteins is one of the most important post-translational modifications of proteins, and reversible phosphorylation controls cellular activities such as cell proliferation, cell division, intercellular information transmission, and gene expression. However, in protein samples, the content of phosphorylated proteins and phosphorylated peptides is very low, and there are interferences from various...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/22G01N1/40
Inventor 訾金张朝良林梁
Owner SHENZHEN HUADA GENE INST
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