Identification of phytochelatin-related peptides in maize seedlings exposed to cadmium and obtained enzymatically in vitro

An analytical strategy based on the sensitivity of electrospray tandem mass spectrometry following a simplified and reproducible sample preparation procedure was evaluated for the determination of Cd-induced phytochelatins (PC) and related peptides in four maize varieties. In addition to the three known families of PC (PC, desGly-PC and iso-PC(Glu)) that were observed, novel PC and desGly-PC homologues lacking the N-terminal ii-linked Glu were isolated from maize root extracts for the first time. Additionally the complete sequence of iso-PC3(Glu) was determined by tandem mass spectrometry. Peptides obtained in vivo and in vitro as the result of the reaction of glutathione with the enzyme phytochelatin synthase were compared. Minor forms detected from in vitro reactions include compounds with intramolecular or intermolecular disulfide bonds resulting from the oxidation of SH groups, phytochelatin homologues lacking the N-terminal gamma -linked Glu, and new PC-related peptides with a Cys-Cys motif. Since peptides lacking a gamma Glu residue could be generated as artifacts in electrospray mass spectrometry, the application of capillary electrophoresis with online electrospray mass spectrometry allowed the separation and detection of such peptides as endogenous molecules present in planta and as products of in vitro reactions. (C) 2001 Elsevier Science Ltd. All rights reserved.