Abstract
We examined the applicability of the “nested” collision induced dissociation/post-source decay (CID/PSD) method to sequencing of novel peptides from solitary wasps which have neurotoxic venom for paralyzing other insects. The CID/PSD spectrum of a ladder peptide derived from an exopeptidase digest was compared with that of the intact peptide. The mass peaks observed only in the CID/PSD spectrum of a ladder peptide were extracted as C-terminal fragment ions. Assignment of C-terminal fragment ions enabled calculation of N-terminal fragment masses, leading to differentiation between N-terminal fragment ions and internal fragment ions. Furthermore, a method incorporating nested CID/PSD combined with endopeptidase digestion is described as an approach to determine the sequence of N-terminally modified peptides. The information from immonium and related ions observed in the CID/PSD spectrum was used for the selection of a suitable endopeptidase for the digestion of peptides. These nested CID/PSD methods allowed rapid and sensitive identification by removing ambiguity in the assignment of the fragment ions, and proved useful for sequencing unknown peptides, in particular those available as natural products with a limited supply.
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