Cloning and expression of a novel Mu class murine glutathione transferase isoenzyme.
The present study describes the cDNA cloning, expression and characterization of a novel Mu class murine glutathione transferase (GST) isoenzyme. Screening of a cDNA library from the small intestine of a female A/J mouse using consensus probes derived from Mu class murine GST genes (mGSTM1-mGSTM5) resulted in the isolation of a full-length cDNA clone of a previously unknown Mu class GST gene (designated as mGSTM7). The choice of tissue was based on our previous identification in female A/J mouse small intestine of a potentially novel Mu class GST isoenzyme. The deduced amino acid sequence of mGSTM7, which comprises of 218 amino acid residues, exhibited about 67-78% identity with other Mu class murine GSTs. Recombinant mGSTM7-7 cross-reacted with anti-(GST Mu) antibodies, but not with anti-(GST Alpha) or anti-(GST Pi) antibodies. The pI and the reverse-phase-HPLC elution profile of recombinant mGSTM7-7 were different from those of other Mu class murine GSTs. The substrate specificity of mGSTM7-7 was also different compared with other Mu class murine GSTs. Interestingly, mGSTM7 had a higher identity with the human Mu class isoenzyme hGSTM4 (87% identity and 94% similarity in the amino acid sequence) than with any of the known mouse Mu class GSTs. Specific activities of recombinant mGSTM7-7 and human GSTM4-4 were comparable towards several substrates. For example, similar to hGSTM4-4, recombinant mGSTM7-7 was poorly active in catalysing the GSH conjugation of 1-chloro-2,4-dinitrobenzene and ethacrynic acid, and lacked activity towards 1,2-dichloro-4-nitrobenzene and 1,2-epoxy-3-(p-nitrophenoxy)propane. These results suggested that hGSTM4-4 might be the human counterpart of mouse GSTM7-7. Reverse transcription-PCR analysis using mGSTM7-specific primers revealed that mGSTM7 is widely expressed in tissues of female A/J mice, including liver, forestomach, lung, kidney, colon and spleen.
Guo
Jianxia
J
Department of Pharmacology and University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, S-871 Scaife Hall (Box 130), PA 15261, USA.
Zimniak
Ludwika
L
Zimniak
Piotr
P
Orchard
John L
JL
Singh
Shivendra V
SV
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D005260:Female; D015723:Gene Library; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000737:chemistry* / Q000235:genetics*; D007421:Intestine, Small / Q000201:enzymology; D051379:Mice; D008969:Molecular Sequence Data; D011487:Protein Conformation; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013379:Substrate Specificity; D013997:Time Factors
D018076:DNA, Complementary; D020033:Protein Isoforms; D012333:RNA, Messenger; D011994:Recombinant Proteins; D004247:DNA; D005982:Glutathione Transferase