Application of Parallel Reaction Monitoring in 15N Labeled Samples for Quantification

Accurate relative quantification is critical in proteomic studies. The incorporation of stable isotope N-15 to plant-expressed proteins in vivo is a powerful tool for accurate quantification with a major advantage of reducing preparative and analytical variabilities. However, N-15 labeling quantification has several challenges. Less identifications are often observed in the heavy-labeled samples because of incomplete labeling, resulting in missing values in reciprocal labeling experiments. Inaccurate quantification can happen when there is contamination from co-eluting peptides or chemical noise in the MS1 survey scan. These drawbacks in quantification can be more pronounced in less abundant but biologically interesting proteins, which often have very few identified peptides. Here, we demonstrate the application of parallel reaction monitoring (PRM) to N-15 labeled samples on a high resolution, high mass accuracy Orbitrap mass spectrometer to achieve reliable quantification even of low abundance proteins in samples.

Automated summary

Accurate relative quantification is crucial in proteomic studies. Incorporating stable isotope N-15 to plant-expressed proteins in vivo is a powerful tool for accurate quantification, but it faces challenges such as incomplete labeling and contamination. This study demonstrates the use of parallel reaction monitoring (PRM) on a high-resolution mass spectrometer to achieve reliable quantification, even for low abundance proteins.