Polypeptide amino acid sequence De novo sequencing method based on chemical modification and isotope labeling

A technology of isotope labeling and chemical modification, applied in the field of de novo sequencing of protein amino acid sequences to identify fragment ions, can solve the problems of affecting the accuracy of de novo sequencing, complex peptide fragmentation mechanism, and reduced probability and intensity, so as to improve specificity, improve Accuracy and efficiency

Active Publication Date: 2017-05-10
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
View PDF5 Cites 7 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, because the fragmentation mechanism of the polypeptide is relatively complicated, even if the polypeptide is subjected to the aforementioned treatment, other fragment ions will still be produced when the polypeptide is fragmented, so the

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Polypeptide amino acid sequence De novo sequencing method based on chemical modification and isotope labeling
  • Polypeptide amino acid sequence De novo sequencing method based on chemical modification and isotope labeling
  • Polypeptide amino acid sequence De novo sequencing method based on chemical modification and isotope labeling

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] 1. Peptide N-terminal labeling and C-terminal labeling based on arginine labeling

[0025] Such as figure 1 As shown, mark according to the following process:

[0026] 1) Guanylation of polypeptide lysine amino groups: Add 40 μL of 100 mM sodium bicarbonate solution containing 2M O-methylisourea to 1 ml of polypeptide solution, adjust the pH value to 11 with 2M sodium hydroxide solution, and incubate at 37 °C React for 2 hours. The reaction was then terminated by adding 10% trifluoroacetic acid and the pH of the solution was adjusted to 8.0.

[0027] 2) Polypeptide N-terminal arginine labeling: 1) the amino group of arginine is blocked with 50 mM sodium phosphate buffer solution (pH 7.5) containing 4% formaldehyde and 60 mM sodium cyanoborohydride; use 1-(3- Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) activate the carboxyl group of arginine; 2) The obtained peptide samples are divided into two equal In the first batch,...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The present invention relates to a polypeptide amino acid sequence De novo sequencing method based on chemical modification and isotope labeling, wherein the MS/MS maps of the same polypeptides having different labels are subjected to correlation by using the correlation between the N-terminal labeling and C-terminal labeling polypeptide molecule quality and the retention time, the polypeptides in different labeled samples form N-terminal fragment ion pair and the C-terminal fragment ion pair during a mass spectrometry fragmentation process, and the polypeptide amino acid sequence is subjected to De novo sequencing. According to the present invention, the polypeptide is modified with the positively charged reagent so as to make the polypeptide easily form the rich fragment ions in the mass spectrometry during fragmentation, and by using the reagents containing different light and weight isotopes to label, the polypeptide form the paired fragment ions in the mass spectrometry during fragmentation so as to be easily distinguished, such that the influence of the interfering signal is reduced, and the fragment ion selection specificity and the De novo sequencing speed can be improved; and by using the complementarity of the N-terminal fragment ions and the C-terminal fragments, the accuracy and the efficiency of the polypeptide sequencing can be improved.

Description

technical field [0001] The present invention relates to a protein amino acid sequence de novo sequencing method assisted by chemical modification and isotope labeling, in particular to a protein amino acid sequence de novo sequencing method for identifying fragment ions by performing stable isotope labeling on the N-terminus and C-terminus of a polypeptide respectively . Background technique [0002] Protein is an important biomacromolecule and plays an important role in life activities. Sequencing proteins is essential for the analysis of protein primary structure. Although there are protein databases for some species, the amino acid sequence of proteins can be analyzed by searching the database. However, most species still do not have protein databases available for searching, and for species that already have databases, the amino acid sequence of their proteins will also change due to genetic variation. Therefore, it is very necessary to develop de novo protein amino a...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): G01N30/02
Inventor 张丽华单亦初张珅陈玲凡张玉奎
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap