Expression of NADP+-dependent 15-hydroxyprostaglandin dehydrogenase mRNA in monkey ocular tissues and characterization of its recombinant enzyme.
Although prostaglandin (PG) F2a and its analogue latanoprost decrease the intraocular pressure in a variety of animals, their intraocular metabolism has not yet been clarified. Here, we isolated the cDNAs for the monkey homologues of NAD+- and NADP+-dependent types of 15-hydroxy PG dehydrogenase (PGDH) from lung and eye, respectively, and investigated the distribution of their mRNAs in the monkey eye. The cDNAs for the NAD+- and NADP+-dependent types of PGDH contained an open reading frame of 798 and 831 bp encoding 266 and 277 amino acid residues with calculated molecular masses of 28.9 and 30.5 kDa, respectively. The amino acid sequences of the monkey NAD+- and NADP+-dependent enzymes showed less than 20% identity to each other, and the former enzyme shows 98.5 and 86.8% identity, and the latter 94.9 and 85.2% identity, to the human and mouse enzymes, respectively. Reverse transcription-PCR analysis revealed that the mRNA for NADP+-dependent PGDH, but not that for NAD+-dependent PGDH, was highly expressed in monkey ocular tissues. In situ hybridization analysis demonstrated that the mRNA for NADP+-dependent PGDH was localized in the epithelial cells of the cornea. The recombinant NADP+-dependent PGDH catalyzed the dehydrogenation of the 15-hydroxyl group of PGF2a and the acid form of latanoprost in the presence of NADP+ as examined by HPLC. These results indicate that PGF2a and the acid form of latanoprost are degraded to their 15-keto metabolites by NADP+-dependent PGDH localized in the monkey eye.
Fujimori
Ko
K
Department of Molecular Behavioral Biology, Japan Science and Technology Corporation, Osaka Bioscience Institute, Furuedai, Suita, Osaka 565-0874, Japan.
Okada
Tetsuya
T
Urade
Yoshihiro
Y
D000595:Amino Acid Sequence; D000818:Animals; D003315:Cornea / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D018076:DNA, Complementary / Q000302:isolation & purification*; D015237:Dinoprost / Q000378:metabolism; D004847:Epithelial Cells / Q000201:enzymology / Q000378:metabolism; D005123:Eye / Q000201:enzymology / Q000378:metabolism*; D000882:Haplorhini; D006801:Humans; D006911:Hydroxyprostaglandin Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D000077338:Latanoprost; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism*; D011461:Prostaglandins F, Synthetic / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D014018:Tissue Distribution
D018076:DNA, Complementary; D011461:Prostaglandins F, Synthetic; D012333:RNA, Messenger; D011994:Recombinant Proteins; D009249:NADP; D000077338:Latanoprost; D015237:Dinoprost; D006911:Hydroxyprostaglandin Dehydrogenases; C035859:15-hydroxyprostaglandin dehydrogenase