DirectMS1Quant: Ultrafast Quantitative Proteomics with MS/MS-Free Mass Spectrometry

Area of Science:

• Proteomics
• Mass Spectrometry
• Quantitative Biology

Background:

• Ultrafast proteome-wide analysis methods are crucial for high-throughput biological studies.
• Existing MS/MS approaches offer high protein identification numbers but may not optimize quantitation quality.
• The DirectMS1 method utilizes minute-long LC gradients and MS1-only spectra for rapid proteome analysis.

Purpose of the Study:

• To compare the quantitation performance of the DirectMS1 method against established MS/MS-based quantitation techniques.
• To evaluate the efficiency and quality of proteome quantitation using MS1-only data analysis.
• To introduce and validate a novel quantitation workflow, DirectMS1Quant.

Main Methods:

• DirectMS1 method with 5 min LC gradients and MS1-only mass spectra acquisition.
• One-on-one comparison with label-free quantitation (LFQ), tandem mass tag (TMT), and data-independent acquisition (DIA) methods.
• Analysis of well-characterized samples, including spiked UPS1 proteins and glioblastoma cell lines.
• Utilized a novel quantitation workflow, DirectMS1Quant, for MS1-only data analysis.

Main Results:

• DirectMS1 achieved a depth of 2500 human cell proteins at 1% FDR with a 7.3 min total runtime.
• The quantitation efficiency of 5 min DirectMS1 was comparable to TMT and DIA methods.
• TMT and DIA methods required 10-20 times longer instrumentation time compared to DirectMS1.

Conclusions:

• DirectMS1 provides a highly efficient and rapid alternative for proteome quantitation.
• MS1-only data analysis can achieve robust quantitation comparable to more time-consuming MS/MS methods.
• The DirectMS1Quant workflow enhances the utility of ultrafast proteome analysis for quantitative studies.