Estimation of genomic complexity, heterologous expression, and enzymatic characterization of mouse glutathione S-transferase mGSTA4-4 (GST 5.7).
We have previously isolated a cDNA clone for a unique mouse lung glutathione S-transferase, mGSTA4-4 (GST 5.7) (Zimniak, P., Eckles, M. A., Saxena, M., and Awasthi, Y. C. (1992) FEBS Lett. 313, 173-176). By genomic Southern blotting and polymerase chain reaction single strand conformation polymorphism analysis we have now demonstrated the presence of at least two mGSTA4-related genes in the mouse. The heterogeneity of mGSTA4-4 was further examined by comparing the structural and kinetic properties of mGSTA4-4 isolated from mouse lung with those of recombinant rec-mGSTA4-4 expressed in Escherichia coli. Except for the isoelectric point, the physical properties of the two proteins were indistinguishable. Western blots using antibodies against rec-mGSTA4-4 have shown selective expression of the enzyme in mouse tissues. Even though the substrate specificity profiles of the tissue-isolated and recombinant enzymes, which point to a role of mGSTA4-4 in the detoxification of lipid peroxidation products, were generally similar, significant differences were observed with selected substrates. The existence of functionally distinct forms of mGSTA4-4 and the presence of more than one gene strongly suggest that the previously observed differences in properties of mGSTA4-4 isolated from various mouse tissues (Awasthi, S., Singhal, S. S., Srivastava, S. K., and Awasthi, Y. C. (1993) Arch. Biochem. Biophys. 301, 143-150) may be due to tissue-specific expression of mGSTA4-related genes.
Zimniak
P
P
Department of Medicine, University of Arkansas for Medical Sciences, Little Rock.
Singhal
S S
SS
Srivastava
S K
SK
Awasthi
S
S
Sharma
R
R
Hayden
J B
JB
Awasthi
Y C
YC
D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D005796:Genes; D005982:Glutathione Transferase / Q000235:genetics*; D007700:Kinetics; D008168:Lung / Q000201:enzymology; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D012150:Polymorphism, Restriction Fragment Length; D012333:RNA, Messenger / Q000235:genetics; D013379:Substrate Specificity; D014018:Tissue Distribution
D017931:DNA Primers; D012333:RNA, Messenger; D005982:Glutathione Transferase