A phi class glutathione S-transferase from Oryza sativa (OsGSTF5): molecular cloning, expression and biochemical characteristics.
A glutathione S-transferase (GST) related to the phi (F) class of enzymes only found in plants has been cloned from the Oryza sativa. The GST cDNA was cloned by PCR using oligonucleotide primers based on the OsGSTF5 (GenBank Accession No. AF309382) sequences. The cDNA was composed of a 669-bp open reading frame encoding for 223 amino acids. The deduced peptide of this gene shared on overall identity of 75% with other known phi class GST sequences. On the other hands, the OsGSTF5 sequence showed only 34% identity with the sequence of the OsGSTF3 cloned by our previous study (Cho et al., 2005). This gene was expressed in Escherichia coli with the pET vector system and the gene product was purified to homogeneity by GSH-Sepharose affinity column chromatography. The expressed OsGSTF5 formed a homo-dimer composed of 28 kDa subunit and its pI value was approximately 7.8. The expressed OsGSTF5 displayed glutathione conjugation activity toward 1-chloro-2,4-dinitrobenzene and 1,2-epoxy-3-(p-nitrophenoxy)propane and glutathione peroxidase activity toward cumene hydroperoxide. The OsGSTF5 also had high activities towards the herbicides alachlor, atrazine and metolachlor. The OsGSTF5 was highly sensitive to inhibition by ShexylGSH, benastatin A and hematin. We propose from these results that the expressed OsGSTF5 is a phi class GST and appears to play a role in the conjugation of herbicide and GPOX activity.
Cho
Hyun-Young
HY
Department of Chemistry, College of Natural Sciences, Chung-Ang University, 221, Huksuk-dong, Dongjak-gu, Seoul 156-756, Korea.
Lee
Hae Joo
HJ
Kong
Kwang-Hoon
KH
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012275:Oryza / Q000201:enzymology*; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity / Q000187:drug effects
D004791:Enzyme Inhibitors; D005982:Glutathione Transferase