DIA-ABPP Target Screening via Label-Free Quantitative Proteomics
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Solution Overview
Problem
Current chemical proteomics techniques, such as isoTOP-ABPP and SLC-ABPP, face challenges with low sensitivity, poor reproducibility, and high costs, limiting their ability to perform large-scale high-throughput detection.
Innovation Solution
The method integrates DIA-based quantitative omics with activity-based protein profiling (ABPP) to form DIA-ABPP, which allows for high coverage, high reproducibility, and high precision target screening without the need for expensive isotope tag reagents.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If isotope tag-based quantitative chemical proteomics techniques (isoTOP-ABPP, SLC-ABPP) are used, then quantitative analysis capability is improved, but cost increases and operability deteriorates
Solution Approach 1:
The patent replaces expensive isotope tag reagents with a cheap and simple label-free quantitative strategy based on DIA mass spectrometry. The method uses direct infusion mass spectrometry without requiring isotopic labeling, making the reagents inexpensive and the procedure more accessible while maintaining quantitative capability through spectral counting and intensity-based approaches.
Solution Approach 2:
The patent extracts and eliminates the complex isotope tagging step from the quantitative chemical proteomics workflow. By removing the need for isotopic labels and their associated complex preparation procedures, the method simplifies the overall process while preserving the core quantitative analysis capability through direct mass spectrometric measurement of protein abundances.
2Measurement precision
If traditional ABPP techniques are used, then target identification capability is improved, but throughput decreases and sensitivity deteriorates
Solution Approach 1:
The patent implements continuous direct infusion mass spectrometry without the intermittent fractionation and sequential analysis required by traditional ABPP methods. The label-free DIA approach allows continuous acquisition of mass spectral data across the entire proteome, enabling high-throughput target identification while maintaining sensitivity through accumulated spectral information and advanced data processing algorithms.
3Measurement precision
If isotope tag reagents are used for quantitative analysis, then quantitation accuracy is improved, but cost increases significantly
Solution Approach 1:
The patent eliminates the use of expensive isotope tag reagents entirely, replacing them with a cost-effective label-free quantitative strategy. The method achieves accurate quantitation through direct measurement of peptide and protein signal intensities in DIA mass spectrometry data, removing the need for costly isotopic labeling reagents while maintaining quantitation accuracy through rigorous data normalization and statistical analysis.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
DIA-ABPP achieves efficient and accurate screening of targetable active amino acid sites, enabling large-scale high-throughput detection and providing valuable insights for drug development.
Implementation Method 1
design an active molecular probe that can covalently bind to certain amino acids in active proteins
Implementation Method 2
quantitative mass spectrometry methods to analyze proteins with specific functions or modifications in complex proteomes
Implementation Method 3
biotin, which affinity enriched by streptavidin resin and used in mass spectrometry to identify proteins bound by active molecular probes
Data Source
AI summary
A method for screening targets by data independent acquisition (DIA)-based quantitative chemical proteomics, comprising covalently modifying specific active amino acids in a proteome with an active molecular probe; and quantitatively analyzing sites covalently-modified by the probe through DIA-based quantitative omics method, to obtain candidate targets. According to the method for screening targets by DIA-based quantitative chemical proteomics, DIA-based quantitative omics technique is applied into activity-based protein profiling (ABPP) to form DIA-ABPP, so that high coverage, high reproducibility and high precision target screening may be achieved, and corresponding technical support is provided for subsequent drug development.