Updated publication reference for PubMed record(s): 29358340. Large-scale N-terminomics using COFRADIC and TAILS has revolutionized protease research, but is disseminated only in few labs. We present an alternative and simple protocol for N-terminal peptide enrichment, based on charge-based fractional diagonal chromatography (ChaFRADIC) and requiring only well-established protein chemistry and a pipette tip. We achieve unprecedented sensitivity quantifying >2000 unique N-terminal peptides from 10 µg per sample, allowing the identification of proteolytic targets and consensus motifs.