Frederike A. Wiebel

Identification of theta-class glutathione S-transferase in liver cytosol of the marmoset monkey.

Abstract The presence of theta-class glutathione S-transferase (GST) in marmoset monkey liver cytosol was investigated. An anti-peptide antibody targeted against the C-terminus of rGSTT1 reacted with a single band in marmoset liver cytosol that corresponded to a molecular weight of 28 kDa. The intensity of the immunoreactive band was not affected by treatment of marmoset monkeys with 2,3,7,8-tetrachlorodibenzo-p-dioxin, phenobarbitone, rifampicin or clofibric acid. Similarly, activity towards methyl chloride (MC) was unaffected by these treatments. However, GST activity towards 1,2-epoxy- 3-(p-nitrophenoxy)-propane (EPNP) was increased in marmosets treated with phenobarbitone (2.6-fold) and rifampicin (2.6-fold), activity towards dichloromethane (DCM) was increased by 50% after treatment of marmosets with clofibric acid, and activity towards 1-chloro-2,4-dinitrobenzene (CDNB) was raised slightly (30–42% increases) after treatment with phenobarbitone, rifampicin or clofibric acid. Compared with humans, marmoset liver cytosol GST activity towards DCM was 18-fold higher, activity towards MC was 7 times higher and activity towards CDNB was 4 times higher. Further, EPNP activity was clearly detectable in marmoset liver cytosol samples, but was undetectable in human samples. Immunoreactive marmoset GST was partially purified by affinity chromatography using hexylglutathione-Sepharose and Orange A resin. The interaction of immunoreactive marmoset GST was similar to that found previously for rat and human GSTT1, suggesting that this protein is also a theta class GST. However, unlike rat GSTT1, the marmoset enzyme was not the major catalyst of EPNP conjugation. Instead, immunoreactivity was closely associated with activity towards MC. In conclusion, these results provide evidence for the presence of theta-class GST in the marmoset monkey orthologous to rGSTT1 and hGSTT1.

Comparison of GST Theta Activity in Liver and Kidney of Four Species

Glutathione transferases (GSTs) catalyzing the conjugation of glutathione with electrophilic substrates are important enzymes in the metabolism of xenobiotics. Several isozymes exhibit polymorphisms in humans. The two deletion polymorphisms of hGSTM1 and hGSTT1 result in total loss of enzyme activity in homozygous null genotype (GSTM1*0 and GSTT1*0 respectively) individuals (Seidegard et al. 1988; Pemble et al. 1994). Individuals that are heterozygous for hGSTT1 show distinctly lower enzyme activities than individuals carrying two functional alleles of hGSTT1 (Wiebel et al. 1996). A similar effect is conceivable for the hGSTM1 polymorphism but has not been verified so far.

Purification and characterization of a new glutathioneS-transferase, class δ, from human erythrocytes

A new polymorphic form of glutathioneS-transferase (GST), metabolising monohalogenated methanes, ethylene oxide and dichloromethane, has been purified from human erythrocytes and characterized. Several characteristics, such as similar elution patterns on different chromatographic matrices, KM-values and activity towards antibodies, confirm a previous assumption that this novel GST is a class δ enzyme. Although the presence or absence of the enzyme activity in human red blood cells is parallel with the polymorphism of the human GST T1 gene, the new GST δ in red blood cells may differ from the known GST T1-1 enzyme from other tissues in terms of substrate specificity, since established GST T1-1 substrates [1,2-epoxy-3-(p-nitro-phenoxy)propane andp-nitrobenzyl chloride] are not metabolized. The substrate specifity of the new enzyme in erythrocytes resembles more closely that of GST T2-2, most likely due to a commonN-terminal modification which modifies substrate binding. The new polymorphic GST-isoform in human red blood cells therefore may be considered to represent anN-terminally modified isoform of GST T1-1.