CYP2R1 (vitamin D 25-hydroxylase) is a microsomal cytochrome P450 monooxygenase, anchored on the cytoplasmic face of the endoplasmic reticulum membrane and expressed mainly in liver (also testis), that catalyses the first step of vitamin D activation: hydroxylation at C-25 of vitamin D3 (calciol) and vitamin D2 to give 25-hydroxyvitamin D (calcidiol), the main circulating vitamin D metabolite and the substrate of renal CYP27B1 1-alpha-hydroxylase. Electrons are supplied by NADPH via cytochrome P450 reductase and the heme iron activates molecular oxygen. Unlike many CYP2 enzymes it has a narrow, secosteroid-specific active site (crystal structure with bound vitamin D3). Biallelic loss-of-function variants (e.g. L99P) cause vitamin D-dependent rickets type 1B with low 25-hydroxyvitamin D; Cyp2r1-null mice have >50% lower serum 25-hydroxyvitamin D, showing CYP2R1 is the major but not the only 25-hydroxylase in vivo.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004497 monooxygenase activity | IEA GO_REF:0000002 | MODIFY | Summary: Generic monooxygenase / paired-donor oxidoreductase parent terms of the specific vitamin D 25-hydroxylase activity. Reason: Correct but uninformative; CYP2R1 has a narrow secosteroid substrate range and its activity is captured precisely by vitamin D 25-hydroxylase activity (GO:0030343). Proposed replacements: vitamin D 25-hydroxylase activity Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt Catalyzes hydroxylation at C-25 of both forms of PMID:18511070 To understand the narrow substrate specificity of CYP2R1 |
| GO:0005506 iron ion binding | IDA PMID:18511070 Structural analysis of CYP2R1 in complex with vitamin D3. | MARK AS OVER ANNOTATED | Summary: Iron ion binding reflects the heme iron of the P450 prosthetic group. Reason: The iron is bound as part of heme, which is captured by heme binding (GO:0020037); free iron ion binding is not a separate activity of the protein. Supporting Evidence: PMID:18511070 Vitamin D3 is bound in an elongated conformation with the aliphatic side-chain pointing toward the heme. |
| GO:0005506 iron ion binding | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: Iron ion binding reflects the heme iron of the P450 prosthetic group. Reason: The iron is bound as part of heme, which is captured by heme binding (GO:0020037); free iron ion binding is not a separate activity of the protein. Supporting Evidence: PMID:18511070 Vitamin D3 is bound in an elongated conformation with the aliphatic side-chain pointing toward the heme. |
| GO:0005737 cytoplasm | IBA GO_REF:0000033 | MODIFY | Summary: Generic cytoplasm location propagated from the CYP2 family. Reason: CYP2R1 is a microsomal enzyme on the endoplasmic reticulum membrane; cytoplasm is too unspecific. Proposed replacements: endoplasmic reticulum membrane Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt SUBCELLULAR LOCATION: Endoplasmic reticulum membrane PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity |
| GO:0005789 endoplasmic reticulum membrane | IEA GO_REF:0000044 | ACCEPT | Summary: CYP2R1 is a microsomal P450 associated with the endoplasmic reticulum membrane. Reason: Microsomal localisation was the basis of its identification as the microsomal 25-hydroxylase (PMID:12867411, PMID:15465040). Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt SUBCELLULAR LOCATION: Endoplasmic reticulum membrane PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-209845 | ACCEPT | Summary: CYP2R1 is a microsomal P450 associated with the endoplasmic reticulum membrane. Reason: Microsomal localisation was the basis of its identification as the microsomal 25-hydroxylase (PMID:12867411, PMID:15465040). Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt SUBCELLULAR LOCATION: Endoplasmic reticulum membrane PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-5602147 | ACCEPT | Summary: CYP2R1 is a microsomal P450 associated with the endoplasmic reticulum membrane. Reason: Microsomal localisation was the basis of its identification as the microsomal 25-hydroxylase (PMID:12867411, PMID:15465040). Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt SUBCELLULAR LOCATION: Endoplasmic reticulum membrane PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity |
| GO:0006766 vitamin metabolic process | TAS Reactome:R-HSA-211916 | MODIFY | Summary: Vitamin metabolic process is a very high-level Reactome-derived term. Reason: The specific process is vitamin D3 biosynthesis (25-hydroxylation step); the broad vitamin term adds no information. Proposed replacements: vitamin D3 biosynthetic process Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt Catalyzes hydroxylation at C-25 of both forms of |
| GO:0006805 xenobiotic metabolic process | IBA GO_REF:0000033 | REMOVE | Summary: Xenobiotic metabolism inferred phylogenetically from the broad CYP2 family. Reason: CYP2R1 is a secosteroid-specific 25-hydroxylase with a narrow active site; there is no evidence that it metabolises xenobiotics physiologically, and the IBA reflects the drug-metabolising function of other CYP2 subfamilies. Supporting Evidence: PMID:18511070 To understand the narrow substrate specificity of CYP2R1 |
| GO:0016705 oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen | IEA GO_REF:0000002 | MODIFY | Summary: Generic monooxygenase / paired-donor oxidoreductase parent terms of the specific vitamin D 25-hydroxylase activity. Reason: Correct but uninformative; CYP2R1 has a narrow secosteroid substrate range and its activity is captured precisely by vitamin D 25-hydroxylase activity (GO:0030343). Proposed replacements: vitamin D 25-hydroxylase activity Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt Catalyzes hydroxylation at C-25 of both forms of PMID:18511070 To understand the narrow substrate specificity of CYP2R1 |
| GO:0016712 oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen, reduced flavin or flavoprotein as one donor, and incorporation of one atom of oxygen | IBA GO_REF:0000033 | MODIFY | Summary: Generic monooxygenase / paired-donor oxidoreductase parent terms of the specific vitamin D 25-hydroxylase activity. Reason: Correct but uninformative; CYP2R1 has a narrow secosteroid substrate range and its activity is captured precisely by vitamin D 25-hydroxylase activity (GO:0030343). Proposed replacements: vitamin D 25-hydroxylase activity Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt Catalyzes hydroxylation at C-25 of both forms of PMID:18511070 To understand the narrow substrate specificity of CYP2R1 |
| GO:0020037 heme binding | IBA GO_REF:0000033 | ACCEPT | Summary: CYP2R1 is a heme-thiolate P450; the heme iron is the catalytic centre and the substrate side chain points toward it in the crystal structure. Reason: Heme binding is directly seen in the crystal structure (PMID:18511070) and is required for the monooxygenase reaction; a supporting (cofactor) function. Supporting Evidence: PMID:18511070 Vitamin D3 is bound in an elongated conformation with the aliphatic side-chain pointing toward the heme. |
| GO:0020037 heme binding | IDA PMID:18511070 Structural analysis of CYP2R1 in complex with vitamin D3. | ACCEPT | Summary: CYP2R1 is a heme-thiolate P450; the heme iron is the catalytic centre and the substrate side chain points toward it in the crystal structure. Reason: Heme binding is directly seen in the crystal structure (PMID:18511070) and is required for the monooxygenase reaction; a supporting (cofactor) function. Supporting Evidence: PMID:18511070 Vitamin D3 is bound in an elongated conformation with the aliphatic side-chain pointing toward the heme. |
| GO:0020037 heme binding | IEA GO_REF:0000002 | ACCEPT | Summary: CYP2R1 is a heme-thiolate P450; the heme iron is the catalytic centre and the substrate side chain points toward it in the crystal structure. Reason: Heme binding is directly seen in the crystal structure (PMID:18511070) and is required for the monooxygenase reaction; a supporting (cofactor) function. Supporting Evidence: PMID:18511070 Vitamin D3 is bound in an elongated conformation with the aliphatic side-chain pointing toward the heme. |
| GO:0030343 vitamin D 25-hydroxylase activity | EXP PMID:12867411 De-orphanization of cytochrome P450 2R1: a microsomal vitami... | ACCEPT | Summary: Vitamin D 25-hydroxylase activity is the defining catalytic activity of CYP2R1, converting vitamin D3/D2 to 25-hydroxyvitamin D. Reason: Established by expression cloning (PMID:12867411), kinetic analysis with purified enzyme (PMID:15465040), the substrate-bound structure (PMID:18511070) and loss of activity in the VDDR1B L99P allele (PMID:15128933). Core function. Supporting Evidence: PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:15128933 eliminated vitamin D 25-hydroxylase enzyme activity |
| GO:0030343 vitamin D 25-hydroxylase activity | IBA GO_REF:0000033 | ACCEPT | Summary: Vitamin D 25-hydroxylase activity is the defining catalytic activity of CYP2R1, converting vitamin D3/D2 to 25-hydroxyvitamin D. Reason: Established by expression cloning (PMID:12867411), kinetic analysis with purified enzyme (PMID:15465040), the substrate-bound structure (PMID:18511070) and loss of activity in the VDDR1B L99P allele (PMID:15128933). Core function. Supporting Evidence: PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:15128933 eliminated vitamin D 25-hydroxylase enzyme activity |
| GO:0030343 vitamin D 25-hydroxylase activity | IDA PMID:15465040 Metabolism of vitamin D by human microsomal CYP2R1. | ACCEPT | Summary: Vitamin D 25-hydroxylase activity is the defining catalytic activity of CYP2R1, converting vitamin D3/D2 to 25-hydroxyvitamin D. Reason: Established by expression cloning (PMID:12867411), kinetic analysis with purified enzyme (PMID:15465040), the substrate-bound structure (PMID:18511070) and loss of activity in the VDDR1B L99P allele (PMID:15128933). Core function. Supporting Evidence: PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:15128933 eliminated vitamin D 25-hydroxylase enzyme activity |
| GO:0030343 vitamin D 25-hydroxylase activity | IDA PMID:18511070 Structural analysis of CYP2R1 in complex with vitamin D3. | ACCEPT | Summary: Vitamin D 25-hydroxylase activity is the defining catalytic activity of CYP2R1, converting vitamin D3/D2 to 25-hydroxyvitamin D. Reason: Established by expression cloning (PMID:12867411), kinetic analysis with purified enzyme (PMID:15465040), the substrate-bound structure (PMID:18511070) and loss of activity in the VDDR1B L99P allele (PMID:15128933). Core function. Supporting Evidence: PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:15128933 eliminated vitamin D 25-hydroxylase enzyme activity |
| GO:0030343 vitamin D 25-hydroxylase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Vitamin D 25-hydroxylase activity is the defining catalytic activity of CYP2R1, converting vitamin D3/D2 to 25-hydroxyvitamin D. Reason: Established by expression cloning (PMID:12867411), kinetic analysis with purified enzyme (PMID:15465040), the substrate-bound structure (PMID:18511070) and loss of activity in the VDDR1B L99P allele (PMID:15128933). Core function. Supporting Evidence: PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:15128933 eliminated vitamin D 25-hydroxylase enzyme activity |
| GO:0030343 vitamin D 25-hydroxylase activity | TAS Reactome:R-HSA-209845 | ACCEPT | Summary: Vitamin D 25-hydroxylase activity is the defining catalytic activity of CYP2R1, converting vitamin D3/D2 to 25-hydroxyvitamin D. Reason: Established by expression cloning (PMID:12867411), kinetic analysis with purified enzyme (PMID:15465040), the substrate-bound structure (PMID:18511070) and loss of activity in the VDDR1B L99P allele (PMID:15128933). Core function. Supporting Evidence: PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:15128933 eliminated vitamin D 25-hydroxylase enzyme activity |
| GO:0030343 vitamin D 25-hydroxylase activity | TAS Reactome:R-HSA-5602147 | ACCEPT | Summary: Vitamin D 25-hydroxylase activity is the defining catalytic activity of CYP2R1, converting vitamin D3/D2 to 25-hydroxyvitamin D. Reason: Established by expression cloning (PMID:12867411), kinetic analysis with purified enzyme (PMID:15465040), the substrate-bound structure (PMID:18511070) and loss of activity in the VDDR1B L99P allele (PMID:15128933). Core function. Supporting Evidence: PMID:12867411 identify an evolutionarily conserved microsomal cytochrome P450 (CYP2R1) with vitamin D 25-hydroxylase activity PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:15128933 eliminated vitamin D 25-hydroxylase enzyme activity |
| GO:0042359 vitamin D metabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: CYP2R1 performs the 25-hydroxylation step of vitamin D activation, part of vitamin D metabolism/biosynthesis of the active hormone. Reason: Correct process assignment supported by the enzyme activity and by reduced 25-hydroxyvitamin D in patients and knockout mice (PMID:15128933, PMID:24019477). Supporting Evidence: PMID:15128933 These data identify CYP2R1 as a biologically relevant vitamin D 25-hydroxylase PMID:24019477 These mice had greater than 50% reduction in serum 25-hydroxyvitamin D3. |
| GO:0042359 vitamin D metabolic process | TAS Reactome:R-HSA-196791 | ACCEPT | Summary: CYP2R1 performs the 25-hydroxylation step of vitamin D activation, part of vitamin D metabolism/biosynthesis of the active hormone. Reason: Correct process assignment supported by the enzyme activity and by reduced 25-hydroxyvitamin D in patients and knockout mice (PMID:15128933, PMID:24019477). Supporting Evidence: PMID:15128933 These data identify CYP2R1 as a biologically relevant vitamin D 25-hydroxylase PMID:24019477 These mice had greater than 50% reduction in serum 25-hydroxyvitamin D3. |
| GO:0042368 vitamin D biosynthetic process | IEA GO_REF:0000041 | ACCEPT | Summary: CYP2R1 performs the 25-hydroxylation step of vitamin D activation, part of vitamin D metabolism/biosynthesis of the active hormone. Reason: Correct process assignment supported by the enzyme activity and by reduced 25-hydroxyvitamin D in patients and knockout mice (PMID:15128933, PMID:24019477). Supporting Evidence: PMID:15128933 These data identify CYP2R1 as a biologically relevant vitamin D 25-hydroxylase PMID:24019477 These mice had greater than 50% reduction in serum 25-hydroxyvitamin D3. |
| GO:0042803 protein homodimerization activity | IDA PMID:18511070 Structural analysis of CYP2R1 in complex with vitamin D3. | KEEP AS NON CORE | Summary: CYP2R1 crystallised as a homodimer. Reason: Homodimer observed in the crystal structure (PMID:18511070); physiological relevance of dimerisation is not established and it is not the core function. Supporting Evidence: file:human/CYP2R1/CYP2R1-uniprot.txt SUBUNIT: Homodimer. |
| GO:1901755 vitamin D3 biosynthetic process | IDA PMID:15465040 Metabolism of vitamin D by human microsomal CYP2R1. | ACCEPT | Summary: CYP2R1 converts vitamin D3 to 25-hydroxyvitamin D3, a step in the formation of hormonally active vitamin D3. Reason: Direct biochemical evidence (PMID:15465040, PMID:18511070); this is the most specific process term for the activity. Supporting Evidence: PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:24019477 CYP2R1 is the major enzyme responsible for 25-hydroxylation of vitamin D, but clearly a second, as-yet unknown, enzyme is another contributor |
| GO:1901755 vitamin D3 biosynthetic process | IDA PMID:18511070 Structural analysis of CYP2R1 in complex with vitamin D3. | ACCEPT | Summary: CYP2R1 converts vitamin D3 to 25-hydroxyvitamin D3, a step in the formation of hormonally active vitamin D3. Reason: Direct biochemical evidence (PMID:15465040, PMID:18511070); this is the most specific process term for the activity. Supporting Evidence: PMID:12867411 indicated that the secosteroid product of CYP2R1 was 25-hydroxyvitamin D3 PMID:15465040 These results strongly suggest that CYP2R1 plays a physiologically important role in the vitamin D 25-hydroxylation in humans. PMID:24019477 CYP2R1 is the major enzyme responsible for 25-hydroxylation of vitamin D, but clearly a second, as-yet unknown, enzyme is another contributor |
| GO:1902271 D3 vitamins binding | IDA PMID:18511070 Structural analysis of CYP2R1 in complex with vitamin D3. | KEEP AS NON CORE | Summary: Vitamin D3 binds in the active site as substrate. Reason: Substrate binding is shown directly in the CYP2R1-vitamin D3 complex structure (PMID:18511070) but is part of the catalytic activity rather than a separate function. Supporting Evidence: PMID:18511070 confirmed the enzyme as a vitamin D 25-hydroxylase, and solved the crystal structure of CYP2R1 in complex with vitamin D3 PMID:18511070 Vitamin D3 is bound in an elongated conformation with the aliphatic side-chain pointing toward the heme. |
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Download this section (compressed HTML)Q: Which enzyme accounts for the residual (~50%) 25-hydroxyvitamin D production in Cyp2r1/Cyp27a1 double-null mice?
Q: Does CYP2R1 contribute to vitamin D activation outside the liver (e.g. testis) in a physiologically meaningful way?
Experiment: Liver-specific versus global Cyp2r1 deletion combined with metabolomic profiling of vitamin D metabolites to locate the residual 25-hydroxylase activity and identify the second enzyme.
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