1. Cellular retinoid-binding proteins transfer retinoids to human cytochrome P450 27C1 for desaturation
- Author
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F. Peter Guengerich and Sarah M. Glass
- Subjects
cytochrome P450 ,Receptors, Retinoic Acid ,medicine.drug_class ,EDC, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride ,Retinoic acid ,AdR, NADPH–adrenodoxin reductase ,retinoid-binding protein ,environment and public health ,Biochemistry ,DNA-binding protein ,vitamin A ,Fatty acid-binding protein ,FABP, fatty acid–binding protein ,ER, endoplasmic reticulum ,Retinoids ,chemistry.chemical_compound ,atROL, all-trans retinol ,CRABP, cellular retinoic acid–binding protein ,enzyme kinetics ,medicine ,Humans ,Ni-NTA, nickel–nitrilotriacetic acid ,Cytochrome P450 Family 27 ,Retinoid ,All trans retinol ,RDH13, retinol dehydrogenase 13 ,Molecular Biology ,Adx, adrenodoxin ,LRAT, lecithin retinol acyltransferase ,biology ,atRAL, all-trans retinaldehyde ,Retinoid binding protein ,CYP or P450, cytochrome P450 ,Cytochrome P450 ,Retinol-Binding Proteins, Cellular ,MS ,atRA, all-trans retinoic acid ,Cell Biology ,Retinoic acid receptor ,protein–protein interaction ,chemistry ,retinoid ,RAR, retinoic acid receptor ,CRBP, cellular retinol-binding protein ,ddRA, 3,4-dehydroretinoic acid ,biology.protein ,dehydroretinoid ,Research Article - Abstract
Cytochrome P450 27C1 (P450 27C1) is a retinoid desaturase expressed in the skin that catalyzes the formation of 3,4-dehydroretinoids from all-trans retinoids. Within the skin, retinoids are important regulators of proliferation and differentiation. In vivo, retinoids are bound to cellular retinol-binding proteins (CRBPs) and cellular retinoic acid–binding proteins (CRABPs). Interaction with these binding proteins is a defining characteristic of physiologically relevant enzymes in retinoid metabolism. Previous studies that characterized the catalytic activity of human P450 27C1 utilized a reconstituted in vitro system with free retinoids. However, it was unknown whether P450 27C1 could directly interact with holo-retinoid-binding proteins to receive all-trans retinoid substrates. To assess this, steady-state kinetic assays were conducted with free all-trans retinoids and holo-CRBP-1, holo-CRABP-1, and holo-CRABP-2. For holo-CRBP-1 and holo-CRABP-2, the kcat/Km values either decreased 5-fold or were equal to the respective free retinoid values. The kcat/Km value for holo-CRABP-1, however, decreased ∼65-fold in comparison with reactions with free all-trans retinoic acid. These results suggest that P450 27C1 directly accepts all-trans retinol and retinaldehyde from CRBP-1 and all-trans retinoic acid from CRABP-2, but not from CRABP-1. A difference in substrate channeling between CRABP-1 and CRABP-2 was also supported by isotope dilution experiments. Analysis of retinoid transfer from holo-CRABPs to P450 27C1 suggests that the decrease in kcat observed in steady-state kinetic assays is due to retinoid transfer becoming rate-limiting in the P450 27C1 catalytic cycle. Overall, these results illustrate that, like the CYP26 enzymes involved in retinoic acid metabolism, P450 27C1 interacts with cellular retinoid-binding proteins.
- Published
- 2021
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