Pseudo-Electropherogram Reconstruction for PTM and Aggregate Analysis
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Solution Overview
Problem
Existing protein mass spectrometry techniques struggle to accurately resolve protein heterogeneity due to post-translational modifications (PTMs) and protein aggregates, especially in complex mixtures, limiting the ability to identify and characterize these forms effectively.
Innovation Solution
Methods and apparatuses for reconstructing a pseudo intact mass spectrum from peptide-level data, allowing for the inclusion or exclusion of specific modified peptide forms, and comparing it with experimentally derived data to identify PTMs and aggregates, using graphical user interfaces for interactive analysis.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If intact protein mass spectrometry is used to capture protein heterogeneity and detect degradation products, then the ability to identify PTMs and aggregates is improved, but measurement precision deteriorates in complex mixtures and when highly abundant species are repeatedly fragmented
Solution Approach 1:
The patent applies segmentation by dividing the intact protein into peptide fragments through proteolytic digestion. This allows the complex protein to be analyzed as smaller, more manageable peptide components that can be precisely identified and characterized, resolving the contradiction between capturing overall protein heterogeneity and achieving precise measurements in complex mixtures.
2Measurement precision
If bottom-up peptide-level analysis is used to achieve highly accurate protein identification, then measurement precision is improved, but the ability to resolve protein heterogeneity due to PTMs and aggregates deteriorates
Solution Approach 1:
The patent merges bottom-up peptide-level analysis with top-down intact protein analysis by integrating peptide mass fingerprinting data with intact protein mass spectrometry data. This combination allows the system to achieve both high measurement precision for protein identification and the ability to resolve protein heterogeneity, including PTMs and aggregates, by correlating peptide-level details with overall protein characteristics.
3Reliability
If peptide-level analysis is used to identify proteins in complex mixtures, then reliability of protein identification is improved, but the ability to detect labile PTMs and provide complete sequence coverage deteriorates
Solution Approach 1:
The patent uses peptide mass fingerprinting as an intermediary approach that bridges intact protein analysis and detailed peptide sequencing. By analyzing peptide masses and patterns without extensive fragmentation, the method reliably identifies proteins in complex mixtures while preserving information about labile PTMs and providing broader sequence coverage compared to traditional MS/MS approaches.
Data Source
AI summary
Methods and apparatuses for the identification and/or characterization of properties of a macromolecule based on mass spectrometry data. Specifically, described herein are methods and apparatuses for converting peptide-level data into a pseudo-intact mass spectra. Also described herein are methods and apparatuses for converting peptide-level data into a pseudo-electropherogram. The methods may be well suited for analyzing proteins and protein complexes, including estimating properties of post-translational modifications of the proteins and protein complexes. Methods may include generating a theoretical graph or spectrum based on peptide-level mass spectrometry data. In some embodiments, the theoretical graph may be a theoretical intact mass spectrum or a theoretical charge distribution spectrum.