Proteomics analysis method based on SP3 enzymolysis

Abstract

The invention aims to provide a proteomics analysis method based on SP3 enzymolysis. The method comprises the following steps: adding lysate into cells of a sample to be detected for denaturation; wherein the component of the lysis solution is 50mM of 4-hydroxyethylpiperazine ethanesulfonic acid; 1% lauryl sodium sulfate, 1% of polyethylene glycol octyl phenyl ether; 1% Tween 20, 1% of ethyl phenyl polyethylene glycol; 5 mM of ethylenediamine tetraacetic acid; 50mM sodium chloride, 1% of glycerol; a 1 * protease inhibitor; and 5 mM dithiothreitol, adding a magnetic bead replacement lysate intothe denatured sample; removing a denaturant influencing trypsin activity; after the replacement is finished, carrying out lysine protease enzymolysis and trypsin enzymolysis on the denatured sample;the method has the advantages that automation can be achieved, the repeatability of experiments of different batches is improved, and meanwhile the flux is improved, and meanwhile the method is not inferior to a traditional protein enzymolysis method in terms of evaluation indexes such as the peptide fragment recovery rate and the sample repeatability.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a proteomics analysis method based on SP3 enzymolysis. Background technique [0002] With the implementation and advancement of the Human Genome Project, life science research has already entered the post-genome era. In this era of greater focus on the function and structure of genes, proteomics has flourished. The concept of proteome (Proteome) was first proposed by Marc Wilkins, which refers to all proteins (Protein) expressed by a genome (genome), or a cell or tissue. Proteomics research mainly includes proteomics qualitative, high-throughput quantitative proteomics, targeted proteomics, and protein post-translational modification. The traditional shotgun method is most widely used in proteomics research. The main process of this method is to hydrolyze protease into peptides by protease, and then analyze it by mass spectrometry, and finally match the...

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Problems solved by technology

[0003] Commonly used proteolysis methods include in-solution enzymatic hydrolysis and Filter-Aided Sample Preparation (abbreviated as FASP). Among them, in-solution enzymatic digestion has restrictions on the composition of the protein solution, and common denaturants (such as SDS, urea, and guanidine hydrochloride) will affect Trypsin activity affects the effect of enzymatic hydrolysis; while FASP enzymatic hydrolysis steps are cumbersome, time-consuming, and sample loss is large, which is not conducive to small samples. Therefore, the invention of new enzymatic hydrolysis methods is of great significance to the development of proteomics

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