| Summary: | The glutathione S-transferase (GSTs) are a multi-gene family of proteins found in all aerobic-life forms; this suggests a fundamental role. These proteins catalyse the nucleophilic attack of the sulphur ion of the tripeptide of glutathione (γ-L-glutamyl-L-cysteine glycine) on a wide range of electrophilic substrates. The conjugation of glutathione to reactive compounds leads ultimately to their detoxification and hence a protective role has been proposed for these enzymes. The majority of the work in this field has focused on the cytosolic GSTs, however a distinct membrane bound GST, the microsomal GST, also exists. Unlike the cytosolic enzymes the microsomal GST holds a latent activity which can be rapidly increased by covalent modification of the protein. The microsomal GST is located in the hydrophobic environment of a number of membranes and it possesses a single cysteine residue which is susceptible to covalent modification by reactive metabolites and oxidised thiol compounds. In order to investigate the role of the microsomal GST in the metabolism of a range of putative substrates an <i>in vivo</i> model was created. The microsomal GST was successfully expressed in <i>Saccharomyces cerevisiae</i> under the control of the PGK promoter. Subcellular fractionation of the yeast demonstrated the microsomal GST to be present in the membrane fraction. In addition the membrane fractions were used to demonstrate the biochemical activity of the expressed protein towards the model GST substrate; 1-chloro-2,4-dinitrobenzene. The results from the cytotoxicity tests, conducted on the yeast model, demonstrated the involvement of the microsomal GST in the metabolism of hexachloro-1,3-butadiene and 1,2-dibromoethane. Hexachloro-1,3-butadiene is a well characterised <i>in vitro</i> substrate for this enzyme, where as 1,2-dibromoethane is not.
|
|---|
