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Characteristic peptide fragment for detecting rat CYP2E1 enzyme and screening method and application thereof

A technology of characteristic peptides and screening methods, which is applied in the field of biological detection and protein, can solve the problems of high quantitative lower limit, insufficient sensitivity, and lack of method verification, and achieve the effect of high sensitivity, strong specificity, and avoiding interference

Active Publication Date: 2018-12-07
余鹏
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method does not use actual samples for verification when screening peptides. It only uses software to screen out specific peptides. Whether it is unique and has a high enough response in the instrument cannot be confirmed in the early screening. At the same time, the current Some methods have not been validated after the method is developed, and the sample quantification is completed only by standard curve, the lower limit of quantification is high, and the sensitivity is not enough

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] This embodiment provides characteristic peptides for detecting rat CYP2E1 enzymes. The characteristic peptides include quantitative peptides, labeled peptides and extended peptides; the quantitative peptides include the first quantitative peptide and the second quantitative peptide. The amino acid sequences of the quantitative peptide and the second quantitative peptide are shown in SEQ ID NO.1-2; the labeled peptide includes the first labeled peptide and the second labeled peptide, the first labeled peptide and the second labeled peptide It is isotope labeling; the extended peptide segment includes a first extended peptide segment and a second extended peptide segment, and the amino acid sequences of the first extended peptide segment and the second extended peptide segment are shown in SEQ ID NO.3-4.

[0046] The isotope labeling method of the first labeled peptide is: FINL( 13 C 6 15 N 1 )VPSNLPHEATR; the isotope labeling mode of the second labeled peptide is GII(...

Embodiment 2

[0049] This embodiment provides a screening method for detecting characteristic peptides of rat CYP2E1 enzymes, comprising the following steps:

[0050] First, according to the amino acid sequence of the CYP2E1 enzyme, the digestion is simulated by Skyline software, and the length of the peptide segment for the simulation digestion is set to be 7-22 amino acids in length. Obtain the simulated enzyme-cleaved peptides, and obtain the ion information of each peptide; then use Uniprot software to exclude regions that do not conform to the characteristic peptides, such as metal binding sites, conflicting sequences, and post-translational modification sites. The Protein BLAST tool of NCBI database (http: / / blast.ncbi.nlm.nih.gov / Blast.cgi) can specifically identify candidate peptides. Input the protein or peptide sequence, select Rattusnorvegicus as the species, and analyze whether the two peptides meet the specificity requirements.

[0051] Secondly, the mouse liver microsome sampl...

Embodiment 3

[0065] This embodiment provides a screening method for detecting characteristic peptides of rat CYP2E1 enzymes, comprising the following steps:

[0066] First, according to the amino acid sequence of the CYP2E1 enzyme, the digestion is simulated by Skyline software, and the length of the peptide segment for the simulation digestion is set to be 7-22 amino acids in length. Obtain the simulated enzyme-cleaved peptides, and obtain the ion information of each peptide; then use Uniprot software to exclude regions that do not conform to the characteristic peptides, such as metal binding sites, conflicting sequences, and post-translational modification sites. The Protein BLAST tool of NCBI database (http: / / blast.ncbi.nlm.nih.gov / Blast.cgi) can specifically identify candidate peptides. Input the protein or peptide sequence, select Rattusnorvegicus as the species, and analyze whether the two peptides meet the specificity requirements.

[0067] Secondly, the mouse liver microsome sampl...

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Abstract

The invention provides a characteristic peptide fragment for detecting a rat CYP2E1 enzyme and a screening method and an application thereof and belongs to the technical field of biodetection and proteins. The characteristic peptide fragment for detecting the rat CYP2E1 enzyme is high in timeliness, can detect the CYP2E1 enzyme specifically and is relatively high in specificity. The peptide fragment can be better prevented from being interfered, the characteristic peptide fragment is applied to detecting the rat CYP2E1 enzyme, and the effect is obvious; the specific fragment can be obtained bymeans of the screening method for the characteristic peptide fragment for detecting the rat CYP2E1 enzyme.

Description

technical field [0001] The invention relates to the field of biological detection and protein technology, in particular to the detection of characteristic peptides of rat CYP2E1 enzymes and its screening method and application. Background technique [0002] Cytochrome P450 (cytochrome P450, CYP450) in the liver is the main enzyme for phase I metabolism of many endogenous and exogenous substances, among which five major enzymes, CYP1A, CYP2C, CYP2D, CYP2E and CYP3A, are involved in 70-80% of the clinically used enzymes. Drug metabolism. CYP2E1 is the metabolic enzyme reported to have the greatest impact on drugs in the CYP2E family. [0003] For drugs that act on CYP2E1 enzymes, different individuals respond differently to the same drug dose, and even lead to unpredictable adverse drug reactions and toxicity, mainly manifested in two aspects: enzyme gene polymorphism and content differences. Its gene polymorphisms have been widely identified, but due to the defects of exist...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/573G01N27/447G01N30/02
CPCG01N27/447G01N30/02G01N33/573G01N33/68
Inventor 余鹏邱朝辉孟凡奇蒋蕾丁尧张可
Owner 余鹏
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