Electrospray ionization-mass spectrometry as a tool for characterization of glutathione S-transferase isozymes

Reversed-phase high-performance liquid chromatography coupled to electrospray ionization-mass spectrometry (ESI-MS) was used for the first time to determine the molecular masses of nine rat liver cytosolic glutathione S-transferase (GST) subunits. The precision of the measurements was +/- 3-4 mass units which, in practice, allowed discrimination between monomers differing by more than 8 Da. Mass accuracy was improved by replicates in the measurements. Comparison of experimental values to cDNA or protein-deduced data available reveals slight differences. N-terminal sequence analyses and interstrain mass comparisons tend to show that primary structures of liver cytosolic GSTs are well conserved from Sprague-Dawley to Wistar rats. Moreover, ESI-MS analysis enabled identification of two minor additional subunits present in both strains, one of which belongs to the mu-class. In addition to rapid and accurate mass determination of GST monomers, and direct determinations achieved on heterodimeric forms, this technique provides precise information on minor structural differences or modifications of these proteins. As such, it constitutes a useful tool for rapid characterization of purified GSTs in comparative studies.