Immunohistochemical localization of guinea-pig leukotriene B4 12-hydroxydehydrogenase/15-ketoprostaglandin 13-reductase.
We have cloned cDNA for leukotriene B4 12-hydroxydehydrogenase (LTB4 12-HD)/15-ketoprostaglandin 13-reductase (PGR) from guinea-pig liver. LTB4 12-HD catalyzes the conversion of LTB4 into 12-keto-LTB4 in the presence of NADP+, and plays an important role in inactivating LTB4. The cDNA contained an ORF of 987 bp that encodes a protein of 329 amino-acid residues with a 78% identity with porcine LTB4 12-HD. The amino acids in the putative NAD+/NADP+ binding domain are well conserved among the pig, guinea-pig, human, rat, and rabbit enzymes. The guinea-pig LTB4 12-HD (gpLTB4 12-HD) was expressed as a glutathione S-transferase (GST) fusion protein in Escherichia coli, which exhibited similar enzyme activities to porcine LTB4 12-HD. We examined the 15-ketoprostaglandin 13-reductase (PGR) activity of recombinant gpLTB4 12-HD, and confirmed that the Kcat of the PGR activity is higher than that of LTB4 12-HD activity by 200-fold. Northern and Western blot analyses revealed that gpLTB4 12-HD/PGR is widely expressed in guinea-pig tissues such as liver, kidney, small intestine, spleen, and stomach. We carried out immunohistochemical analyses of this enzyme in various guinea-pig tissues. Epithelial cells of calyx and collecting tubules in kidney, epithelial cells of airway, alveoli, epithelial cells in small intestine and stomach, and hepatocytes were found to express the enzyme. These findings will lead to the identification of the unrevealed roles of PGs and LTs in these tissues.
Yamamoto
T
T
The Department of Biochemistry and Molecular Biology, CREST of Japan Science and Technology Corporation, Tokyo, Japan.
Yokomizo
T
T
Nakao
A
A
Izumi
T
T
Shimizu
T
T
D011452:15-Oxoprostaglandin 13-Reductase / Q000235:genetics / Q000378:metabolism*; D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D006168:Guinea Pigs; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution
D018076:DNA, Complementary; D012333:RNA, Messenger; D000429:Alcohol Oxidoreductases; C082646:leukotriene B4 12-hydroxydehydrogenase; D011452:15-Oxoprostaglandin 13-Reductase