Cleavable Mass Tags for Accurate Peptide Quantification
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Solution Overview
Problem
Existing isobaric labeling methods for mass spectrometry suffer from interference by co-isolation and co-fragmentation of peptide ions, leading to decreased accuracy and precision in quantification and identification, particularly for peptides with higher charge states and high-mobility protons.
Innovation Solution
A compound with a reactive moiety that forms a covalent bond with a peptide and a moiety that fragments at lower energies than peptides, allowing for selective fragmentation of the compound while leaving the peptide intact, thereby enhancing quantification and identification accuracy.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If mass spectrometry is operated at high energy to fragment peptides for identification, then peptide identification is enabled, but quantification accuracy deteriorates due to co-fragmentation of interfering peptide ions
Solution Approach 1:
The mass tag is divided into two functional segments: a reporter moiety for quantification and a balance moiety for mass balancing. The cleavable bond between these segments allows selective fragmentation of the tag at lower energies without requiring full peptide backbone fragmentation, thereby resolving the contradiction between quantification accuracy and device complexity
Solution Approach 2:
A cleavable bond with specific fragmentation energy serves as an intermediary mechanism that enables preferential fragmentation of the mass tag over the peptide backbone. This intermediary bond allows the system to achieve quantification at lower energies before peptide fragmentation occurs, improving quantification accuracy while reducing the energy complexity required
2Measurement precision
If additional gas-phase isolation steps are performed to address co-isolation issues, then quantification accuracy improves, but acquisition speed deteriorates
Solution Approach 1:
The mass tag is designed with a cleavable bond that fragments at a predetermined lower energy level before peptide backbone fragmentation occurs. This preliminary fragmentation action separates the quantification signal from interfering peptide ions, improving quantification accuracy without requiring additional isolation steps that would slow down acquisition
3Measurement precision
If MS3 scans are performed to improve quantification, then accuracy improves, but sensitivity deteriorates
Solution Approach 1:
The reporter moiety is extracted as a separate fragment from the mass tag through selective cleavage at the cleavable bond. This extracted reporter signal can be quantified directly without requiring MS3 scans, improving sensitivity while maintaining accuracy by eliminating the need for additional scanning steps
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
The compound achieves higher conversion rates of fragmentation compared to peptides, enabling precise quantification and identification by disentangling quantification from peptide fragmentation, and can be used for MS-based enrichment of peptides of interest.
Implementation Method 1
a reactive moiety, said reactive moiety being capable of reacting with a functional group of a peptide to form a covalent bond
Implementation Method 2
a moiety which fragments in the mass spectrometer at an energy below the energy required for fragmenting peptides
Data Source
AI summary
The present invention relates to a compound which comprises or consists of (a) a reactive moiety, said reactive moiety being capable of reacting with a functional group of a peptide to form a covalent bond; and covalently joined thereto (b) a moiety which fragments in the mass spectrometer (i) at an energy below the energy required for fragmenting peptides and/or a higher conversion rate than peptides; and (ii) at said energy according to (i) and when coupled to a peptide via said reactive group, at a single site within said compound coupled to a peptide, to yield a first moiety and a second moiety, said second moiety being coupled to said peptide.