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Enhanced Discovery of Alternative Proteins (AltProts) in Mouse Cardiac Development Using Data-Independent Acquisition
Yuanliang Zhang1, Ying Yang1, Kecheng Li1
1Department of Applied Biology and Chemical Technology, State Key Laboratory of Chemical Biology and Drug Discovery, Hong Kong Polytechnic University, Hong Kong 999077, China.
Analytical Chemistry
|January 15, 2025
Summary
This study introduces a new mass spectrometry (MS) method for detecting alternative proteins (AltProts), increasing identification by two-fold. The optimized workflow enhances AltProt discovery and analysis, revealing new proteins involved in heart development.
Area of Science:
- Proteomics
- Molecular Biology
- Biochemistry
Background:
- Alternative proteins (AltProts) are encoded by noncoding DNA sequences and have distinct biological roles.
- Data-independent acquisition (DIA) mass spectrometry (MS) is underutilized for AltProt research due to library construction challenges.
Purpose of the Study:
- To develop a novel DIA-MS workflow for efficient AltProt detection.
- To establish an optimal AltProt analysis pipeline by comparing databases, library strategies, and software.
- To investigate AltProt expression during mouse heart development.
Main Methods:
- A fragmentation spectra predictor was used to create DIA libraries for AltProt detection.
- Comparative analysis of AltProt databases, DIA library construction strategies, and analytical software.
- Application of the optimized workflow to analyze differential AltProt expression in mouse embryonic and adult heart tissues.
Main Results:
- The novel DIA workflow significantly enhanced AltProt identification (2-fold increase) and reduced missing values (50%) compared to data-dependent acquisition (DDA).
- Over 50 differentially expressed AltProts were identified during mouse heart development.
- More than 30 unannotated mouse AltProts were validated, including ASDURF, crucial for cardiac development.
Conclusions:
- A practical and efficient DIA-MS workflow for AltProt analysis has been established.
- The study reveals novel AltProts with potential roles in biological functions, particularly in cardiac development.
