Identification of mu-class glutathione transferases M2-2 and M3-3 as cytosolic prostaglandin E synthases in the human brain.
Cytosolic prostaglandin (PG) E synthase was purified from human brain cortex. The N-terminal amino acid sequence, PMTLGYXNIRGL, was identical to that of the human mu-class glutathione transferase (GST) M2 subunit. Complementary DNAs for human GSTM2, GSTM3, and GSTM4 subunits were cloned, and recombinant proteins were expressed as homodimers in Escherichia coli. The recombinant GSTM2-2 and 3-3 catalyzed the conversion of PGH2 to PGE2 at the rates of 282 and 923 nmol/min/mg of protein, respectively, at the optimal pH of 8, whereas GSTM4-4 was inactive; although all three enzymes showed GST activity. The PGE synthase activity depended on thiols, such as glutathione, dithiothreitol, 2-mercaptoethanol, or L-cysteine. Michaelis-Menten constants and turnover numbers for PGH2 were 141 microM and 10.8 min(-1) for GSTM2-2 and 1.5 mM and 130 min(-1) for GSTM3-3, respectively. GSTM2-2 and 3-3 may play crucial roles in temperature regulation, nociception, and sleep-wake regulation by producing PGE2 in the brain.
Beuckmann
C T
CT
Department of Molecular Behavioral Biology, Osaka Bioscience Institute, Suita, Japan.
Fujimori
K
K
Urade
Y
Y
Hayaishi
O
O
D000595:Amino Acid Sequence; D002540:Cerebral Cortex / Q000201:enzymology*; D003001:Cloning, Molecular; D003600:Cytosol / Q000201:enzymology; D005982:Glutathione Transferase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D019746:Intramolecular Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D010446:Peptide Fragments / Q000737:chemistry; D000071876:Prostaglandin-E Synthases; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013438:Sulfhydryl Compounds / Q000378:metabolism
D007527:Isoenzymes; D010446:Peptide Fragments; D021122:Protein Subunits; D011994:Recombinant Proteins; D013438:Sulfhydryl Compounds; D005982:Glutathione Transferase; C412602:glutathione S-transferase Mu 2; C416969:glutathione transferase M3-3; D019746:Intramolecular Oxidoreductases; D000071876:Prostaglandin-E Synthases