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An electrophoretic profiling method for thiol‐rich phytochelatins and metallothioneins
Author(s) -
Fan Teresa WM.,
Lane Andrew N.,
Higashi Richard M.
Publication year - 2004
Publication title -
phytochemical analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 72
eISSN - 1099-1565
pISSN - 0958-0344
DOI - 10.1002/pca.765
Subject(s) - chemistry , thiol , chromatography , electrophoresis , capillary electrophoresis , chelation , combinatorial chemistry , biochemistry , inorganic chemistry
Abstract Thiol‐rich peptides such as phytochelatins (PCs) and metallothioneins (MTs) are important cellular chelating agents which function in metal detoxication and/or homeostasis. The variations in molecular sizes and lack of chromophores of these peptides make their analysis difcult. This paper reports an electrophoresis‐based method for a broad screen of thiol‐rich peptides and proteins. The method uses the thiol‐selective uorescent tag, monobromobimane, coupled with Tricine–sodium dodecyl sulphate–urea polyacrylamide gel electrophoresis for a sensitive determination of both PCs and MTs. Results for PCs were conrmed by two‐dimensional NMR and HPLC‐tandem MS analyses. Sample throughput is substantially improved over chromatography‐based methods through parallel sample analysis in 1 h of electrophoretic separation. The method is versatile in that peptides ranging from glutathione to large proteins can be analysed by simple modication(s) of the extraction and electrophoretic conditions, and the nature of the method supports serendipitous detection of unexpected or novel thiol metabolites. Copyright © 2004 John Wiley & Sons, Ltd.