Enzyme and Chemical Assisted N-Terminal Blocked Peptides Analysis, ENCHANT, as a Selective Proteomics Approach...

Shotgun (bottom-up) approach has been widely applied in large-scale proteomics studies. The inherent shortages of shotgun approach lie in that the generated peptides often overwhelm the analytical capacity of current LC-MS/MS systems and that high-abundance proteins often hamper the identification of low-abundance proteins when analyzing complex samples. To reduce the sample complexity and relieve the problems caused by abundant proteins, herein we introduce a modified selective proteomics approach, termed ENCHANT, for enzyme and chemical assisted N-terminal blocked peptides analysis. Modified from our previous Nα-acetylome approach, ENCHANT aims to analyze three kinds of peptides, acetylated protein N-termini, N-terminal glutamine and N-terminal cysteine containing peptides. Application of ENCHANT to HeLa cells allowed to identify 3375 proteins, 19.6% more than that by conventional shotgun approach. More importantly, ENCHANT demonstrated an excellent complementarity to conventional shotgun approach with the overlap of 34.5%. In terms of quantification using data independent acquisition (DIA) technology, ENCHANT quantified 23.9% more proteins than conventional shotgun approach with the overlap of 27.6%. Therefore, our results strongly suggest that ENCHANT is a promising selective proteomics approach, which is complementary to conventional shotgun approach in both qualitative and quantitative proteomics studies. Data are available via ProteomeXchange with identifier PXD007863.