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Due to mechanisms such as proteolytic processing or alternative translation starts, in vivo proteoforms do not necessarily correspond directly to those encoded in the genome. Therefore, the knowledge of protein termini is an indispensable prerequisite to understand protein functions. So far, sequencing of protein N- and C-termini has been limited to single purified protein species, while the proteome-wide identification of N- and C-termini relies on the generation of single, terminal proteotypic peptides followed by chemical enrichment