Determination of SRM peptide transitions by multiplexed product-ion scan mode

Abstract

Selected reaction monitoring (SRM) is a targeted proteomics approach that provides high through-put, sensitive and accurate quantification of proteins in complex biological samples. A key prerequisite to successful SRM assay is precise selection and optimization of SRM peptide transitions of target proteins of interest. SRM peptide transitions can be determined based on their MS/MS spectra utilizing proteomics profiling data or various SRM peptide transition prediction tools. Although in-silico predictions for the SRM peptide transitions is most commonly used approach, reliable SRM peptide transitions selected from actual experiment are desired. Here we demonstrated two modes of Triple Quadrupole masss pectrometry (QqQ-MS) todetermine SRM peptide transitions using peptide profiling data information acquired from Linear Ion-Trap mass spectrometry (LIT-MS). These approaches outperform determination of SRM peptide transitions in a multiplexing manner and can complement to the widely used in-silico SRM peptide transition prediction method. With the predetermined SRM peptide transitions of key oxidative phosphorylation (OXPHOS) proteins in mitochondria, we performed SRM assay to demonstrate the reliability of the SRM peptide transitions for the detection of target peptides in biological sample matrices in human skeletal muscle tissue sample.