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 sheep blowfly) pupae were purified by affinity chromatography and anion-exchange chromatography. One isoenzyme was composed of M(r)-24,800 subunits, and two isoenzymes had subunits of M(r) 23,900. The M(r)-23,900 subunits showed immunological identity and were immunologically distinct from the M(r)-24,800 subunits. All three enzymes were active with the substrate 1-chloro-2,4-dinitrobenzene and had low activity with 1,2-dichloro-4-nitrobenzene. A cDNA clone encoding a M(r)-23,900 subunit (LuGST1) was isolated and sequenced. The sequence has close similarities (> 81%) to that of GSTs from the fruitfly Drosophila melanogaster and Musca domestica (housefly). The deduced amino acid sequence of the Lu GST1 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 identified many hybridizing fragments. Taken together, these data indicated that the L. cuprina genome contains multiple glutathione S-transferase genes.
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Molecular Genetics Group, John Curtin School of Medical Research, Australian National University, Canberra.
Russell
R J
RJ
Marano
R J
RJ
Oakeshott
J G
JG
D000595:Amino Acid Sequence; D000818:Animals; D001315:Australia; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004175:Diptera / Q000201:enzymology* / Q000235:genetics; D004331:Drosophila melanogaster / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D005982:Glutathione Transferase / Q000096:biosynthesis* / Q000302:isolation & purification* / Q000378:metabolism; D006793:Houseflies / Q000201:enzymology; D007527:Isoenzymes / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid; D012756:Sheep; D013379:Substrate Specificity
D007527:Isoenzymes; D046911:Macromolecular Substances; D005982:Glutathione Transferase