Influence of reducing agents on the integrity of selenocompounds. Exploratory work for selenoproteome analysis

Abstract

Three selenium species (selenomethionine, selenocystine and selenocystamine) were studied for their behaviour in the presence of the reducing agents dithiothreitol and tributylphosphine, and of a derivatising agent, iodoacetic acid. These chemicals are commonly used in proteome analysis by means of gel electrophoresis. To study the effect of these chemicals on the selenospecies, two separation techniques were used, capillary electrophoresis and liquid chromatography, both with ICP-MS as an element-specific detector. Electrospray-mass spectrometry provided molecular information. Tributylphosphine (TBT) was shown to partly reduce selenocystine to selenocysteine, and iodoacetic acid to derivatise all the species, with conservation of the species information. Dithiothreitol (DTT), on the contrary, partly sequestered selenium from the species to form a DTT–Se molecule.