Characterization of a novel porcine carbonyl reductase activated by glutathione: Relationship to carbonyl reductase 1, 3α/β-hydroxysteroid dehydrogenase and prostaglandin 9-ketoreductase.

A porcine gene, LOC100622246, encodes carbonyl reductase [NADPH] 1 (pCBR-N1), whose function remains unknown. Previously, three porcine carbonyl reductases, carbonyl reductase 1 (pCBR1), 3α/β-hydroxysteroid dehydrogenase (p3α/β-HSD) and prostaglandine-9-keto reductase (pPG-9-KR), were purified from neonatal testis, adult testis and adult kidney, respectively. However, the relationship of pCBR-N1 with the three enzymes is still unknown. Here, we compare the properties of the recombinant pCBR-N1 and pCBR1. The two enzymes reduced various carbonyl compounds including 5α-dihydrotestosterone, which was converted to its 3α- and 3β-hydroxy-metabolites. Compared to pCBR1, pCBR-N1 exhibited higher Km and kcat values for most substrates, but more efficiently reduced prostaglandin E2. pCBR-N1 was inhibited by known inhibitors of p3α/β-HSD (hexestrol and indomethacin), but not by pCBR1 inhibitors. pCBR-N1 was highly expressed than pCBR1 in the several tissues of adult domestic and microminiature pigs. The results, together with partial amino acid sequence match between pCBR-N1 and pPG-9-KR, reveal that pCBR-N1 is identical to p3α/β-HSD and pPG-9-KR. Notably, pCBR-N1, but not pCBR1, reduced S-nitrosoglutathione and glutathione-adducts of alkenals including 4-oxo-2-nonenal with Km of 8.3-32 μM, and its activity toward non-glutathionylated substrates was activated 2- to 9-fold by 1 mM glutathione. Similar activation by glutathione was also observed for human CBR1. Site-directed mutagenesis revealed that the differences in kinetic constants and glutathione-mediated activation between pCBR-N1 and pCBR1 are due to differences in residue 236 and two glutathione-binding residues (at positions 97 and 193), respectively. Thus, pCBR-N1 is a glutathione-activated carbonyl reductase that functions in the metabolism of endogenous and xenobiotic carbonyl compounds. Copyright © 2023 Elsevier B.V. All rights reserved.

Citation

Satoshi Endo, Yoshifumi Morikawa, Toshiyuki Matsunaga, Akira Hara, Masaki Takasu. Characterization of a novel porcine carbonyl reductase activated by glutathione: Relationship to carbonyl reductase 1, 3α/β-hydroxysteroid dehydrogenase and prostaglandin 9-ketoreductase. Chemico-biological interactions. 2023 Aug 25;381:110572

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PMID: 37247810

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