Identification and expression of a novel class of glutathione -S -transferase from amphioxus Branchiostoma belcheri with implications to the origin of vertebrate liver

Abstract Glutathione- S -transferases have been identified in all the living species examined so far, yet little is known to date about them in amphioxus, a model organism for insights into the origin and evolution of vertebrates. We have isolated a cDNA encoding an amphioxus ( Branchiostoma belcheri ) glutathione- S -transferase with a predicted molecular mass of approximately 26 kDa, from the gut cDNA library. The glutathione- S -transferase had 43.7–51.8% identity to most glutathione- S -transferases identified from aquatic organisms including fish and green alga, but it was much less identical ( S -transferase classes. The phylogenetic analysis revealed that the glutathione- S -transferase was grouped together with most piscine and algal glutathione- S -transferases, separating from other cytosolic glutathione- S -transferase classes. Moreover, the glutathione- S -transferase had an exon–intron organization typical of zebrafish putative GST, red sea bream GSTR1 and plaice GSTA1 genes. The recombinant glutathione- S -transferase has been successfully expressed and purified, which showed a relatively high catalytic activity (3.37 ± 0.1 unit/mg) toward 1-chloro-2, 4-dinitrobenzene and a moderate activity toward ethacrynic acid (0.41 ± 0.01 unit/mg), although it had no detectable activity toward 1, 2-dichloro-4-nitrobenzene, 4-hydroxynonenal, 4-nitrobenzyl chloride and cumene hydroperoxide. In addition, we have revealed a tissue-specific expression pattern of the glutathione- S -transferase gene in B. belcheri , with the most abundant expression in the hepatic caecum. All these indicate that the amphioxus glutathione- S -transferase belongs to a novel rho-class of glutathione- S -transferases with a tissue-specific expression pattern. The relation between the glutathione- S -transferase expression in amphioxus hepatic caecum and the origin of vertebrate liver is also discussed.