PURIFICATION, MOLECULAR-CLONING AND HETEROLOGOUS EXPRESSION OF A GLUTATHIONE-S-TRANSFERASE FROM THE AUSTRALIAN SHEEP BLOWFLY (LUCILIA-CUPRINA)

Three glutathione S-transferases from Lucilia cuprina (Australian shee p blowfly) pupae were purified by affinity chromatography and anion-ex change chromatography. One isoenzyme was composed of M(r)-24800 subuni ts, and two isoenzymes had subunits of M(r) 23900. The M(r)-23900 subu nits showed immunological identity and were immunologically distinct f rom the M(r)-24800 subunits. All three enzymes were active with the su bstrate -1chloro-2,4-dinitrobenzene and had low activity with 1,2-dich loro-4-nitrobenzene. A cDNA clone encoding a M(r)-23 900 subunit (LuGS T1) was isolated and sequenced. The sequence has close similarities (> 81 %) to that of GSTs from the fruitfly Drosophila melanogaster and M usca domestica (housefly). The deduced amino acid sequence of the Lu G ST1 subunit showed no significant similarity to that of the mammalian GSTs to the Alpha, Mu and Pi classes, but shows some similarity (33 %) over the first 100 residues with the rat subunit 12 Theta-class GST. Southern blots of genomic DNA hybridized with the LuGST1 cDNA identifi ed many hybridizing fragments. Taken together, these data indicated th at the L. cuprina genome contains multiple glutathione S-transferase g enes.