{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,20]],"date-time":"2026-10-20T12:44:51Z","timestamp":1792500291110,"version":"4.5.0"},"reference-count":35,"publisher":"Wiley","issue":"3","license":[{"start":{"date-parts":[[2025,10,5]],"date-time":"2025-10-05T00:00:00Z","timestamp":1759622400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2025,10,5]],"date-time":"2025-10-05T00:00:00Z","timestamp":1759622400000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"funder":[{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["19K05829"],"award-info":[{"award-number":["19K05829"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001691","name":"Japan Society for the Promotion of Science","doi-asserted-by":"publisher","award":["21380067"],"award-info":[{"award-number":["21380067"]}],"id":[{"id":"10.13039\/501100001691","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["febs.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["The FEBS Journal"],"published-print":{"date-parts":[[2026,2]]},"abstract":"<jats:p>\n                    The cytochrome P450 3A4 (CYP3A4) variant I301T has been associated with type 3 rickets, which is characterized by reduced serum calcium and significantly decreased levels of 25\u2010hydroxyvitamin D3 [25(OH)D3] and 1\u03b1,25\u2010dihydroxyvitamin D3 [1\u03b1,25(OH)\n                    <jats:sub>2<\/jats:sub>\n                    D3]. Although enhanced 4\u2010hydroxylation of these metabolites was previously proposed as the underlying mechanism causing the disease, the precise enzymatic basis remained unclear. In this study, we investigated the enzymatic properties of CYP3A4\u2010I301T using membrane fractions from recombinant\n                    <jats:italic>Escherichia coli<\/jats:italic>\n                    expressing the variant. Surprisingly, we found that CYP3A4\u2010I301T efficiently produced additional metabolites compared to the wild\u2010type CYP3A4, including 11\u03b1,25\u2010dihydroxyvitamin D3 and 1\u03b1,11\u03b1,25\u2010trihydroxyvitamin D3, which have much lower affinity for the vitamin D receptor than do 25(OH)D3 and 1\u03b1,25(OH)\n                    <jats:sub>2<\/jats:sub>\n                    D3, respectively. Docking simulations suggested that the 3\u03b2\u2010hydroxyl group of 25(OH)D3 forms a hydrogen bond with Thr301 of CYP3A4\u2010I301T, while the 25\u2010hydroxyl group interacts with Arg372 and Glu374, favoring hydroxylation at the 11\u03b1\u2010position. These results indicate that the I301T mutation confers a gain\u2010of\u2010function phenotype on CYP3A4, enhancing its ability to metabolize 25(OH)D3 and 1\u03b1,25(OH)\n                    <jats:sub>2<\/jats:sub>\n                    D3 into low\u2010activity derivatives. This enzymatic shift likely contributes to substrate depletion and impaired calcium homeostasis in affected individuals. Our findings provide new mechanistic insight into the pathological consequences of a single amino acid substitution in CYP3A4 and expand the understanding of enzyme gain\u2010of\u2010function mutations in metabolic disorders.\n                  <\/jats:p>","DOI":"10.1111\/febs.70277","type":"journal-article","created":{"date-parts":[[2025,10,5]],"date-time":"2025-10-05T09:55:00Z","timestamp":1759658100000},"page":"660-676","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Distinct and potent vitamin D hydroxylation activity acquired by the\n                    <scp>CYP3A4 I301T<\/scp>\n                    single amino acid substitution causes type 3 rickets"],"prefix":"10.1111","volume":"293","author":[{"given":"Naoto","family":"Nakaya","sequence":"first","affiliation":[{"name":"Department of Pharmaceutical Engineering Faculty of Engineering, Toyama Prefectural University  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ryota","family":"Sakamoto","sequence":"additional","affiliation":[{"name":"Department of Biotechnology and Life Science Faculty of Engineering, Tokyo University of Agriculture and Technology  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hiroki","family":"Mano","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Engineering Faculty of Engineering, Toyama Prefectural University  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bunzo","family":"Mikami","sequence":"additional","affiliation":[{"name":"Research Institute for Sustainable Humanosphere Kyoto University  Japan"},{"name":"Institute of Advanced Energy Kyoto University  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hiromasa","family":"Imaishi","sequence":"additional","affiliation":[{"name":"Division of Signal Responses, Biosignal Research Center Kobe University  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kazuo","family":"Nagasawa","sequence":"additional","affiliation":[{"name":"Department of Biotechnology and Life Science Faculty of Engineering, Tokyo University of Agriculture and Technology  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7064-8185","authenticated-orcid":false,"given":"Toshiyuki","family":"Sakaki","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Engineering Faculty of Engineering, Toyama Prefectural University  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3495-3216","authenticated-orcid":false,"given":"Kaori","family":"Yasuda","sequence":"additional","affiliation":[{"name":"Department of Pharmaceutical Engineering Faculty of Engineering, Toyama Prefectural University  Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2025,10,5]]},"reference":[{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.abb.2011.11.003"},{"key":"e_1_2_9_3_1","doi-asserted-by":"publisher","DOI":"10.2741\/1514"},{"key":"e_1_2_9_4_1","doi-asserted-by":"publisher","DOI":"10.1111\/febs.12862"},{"key":"e_1_2_9_5_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jsbmb.2016.07.014"},{"key":"e_1_2_9_6_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00223-020-00784-2"},{"key":"e_1_2_9_7_1","first-page":"73","article-title":"Molecular genetics of vitamin D\u2010 dependent hereditary rickets","volume":"57","author":"Kato S","year":"2002","journal-title":"Horm Res"},{"key":"e_1_2_9_8_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M100898200"},{"key":"e_1_2_9_9_1","doi-asserted-by":"publisher","DOI":"10.1124\/mol.111.076356"},{"key":"e_1_2_9_10_1","doi-asserted-by":"publisher","DOI":"10.1124\/mol.105.017392"},{"key":"e_1_2_9_11_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2021.100668"},{"key":"e_1_2_9_12_1","doi-asserted-by":"publisher","DOI":"10.1590\/S0004-282X2004000600003"},{"key":"e_1_2_9_13_1","doi-asserted-by":"publisher","DOI":"10.1210\/jc.2016-4048"},{"key":"e_1_2_9_14_1","doi-asserted-by":"publisher","DOI":"10.1172\/JCI98680"},{"key":"e_1_2_9_15_1","article-title":"CYP3A4 mutation causes vitamin D\u2010dependent rickets type 3: a case report in Saudi Arabia","volume":"15","author":"Al\u2010Ashwal A","year":"2023","journal-title":"Cureus"},{"key":"e_1_2_9_16_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00223-022-01051-2"},{"key":"e_1_2_9_17_1","doi-asserted-by":"publisher","DOI":"10.3390\/biom13071036"},{"key":"e_1_2_9_18_1","doi-asserted-by":"publisher","DOI":"10.1002\/jbmr.1839"},{"key":"e_1_2_9_19_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0021-9258(20)82244-3"},{"key":"e_1_2_9_20_1","doi-asserted-by":"publisher","DOI":"10.1359\/JBMR.0301257"},{"key":"e_1_2_9_21_1","doi-asserted-by":"publisher","DOI":"10.3390\/biom14050551"},{"key":"e_1_2_9_22_1","doi-asserted-by":"publisher","DOI":"10.1046\/j.0014-2956.2001.02615.x"},{"key":"e_1_2_9_23_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.atherosclerosis.2014.06.005"},{"key":"e_1_2_9_24_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.pharmthera.2012.12.007"},{"key":"e_1_2_9_25_1","doi-asserted-by":"publisher","DOI":"10.1210\/jc.2004-0966"},{"key":"e_1_2_9_26_1","doi-asserted-by":"publisher","DOI":"10.1002\/cbic.202100250"},{"key":"e_1_2_9_27_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbrc.2004.08.110"},{"key":"e_1_2_9_28_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbrc.2004.07.040"},{"key":"e_1_2_9_29_1","doi-asserted-by":"publisher","DOI":"10.1021\/ol061955a"},{"key":"e_1_2_9_30_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bmcl.2017.07.060"},{"key":"e_1_2_9_31_1","doi-asserted-by":"publisher","DOI":"10.1021\/jo00157a004"},{"key":"e_1_2_9_32_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0040-4020(01)89344-3"},{"key":"e_1_2_9_33_1","doi-asserted-by":"publisher","DOI":"10.1021\/ac103059k"},{"key":"e_1_2_9_34_1","doi-asserted-by":"publisher","DOI":"10.4014\/jmb.2003.03032"},{"key":"e_1_2_9_35_1","doi-asserted-by":"publisher","DOI":"10.1248\/cpb.c23-00395"},{"key":"e_1_2_9_36_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jsbmb.2017.10.024"}],"updated-by":[{"DOI":"10.1111\/febs.70771","type":"correction","label":"Correction","source":"publisher","updated":{"date-parts":[[2026,10,20]],"date-time":"2026-10-20T00:00:00Z","timestamp":1792454400000}}],"container-title":["The FEBS Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/febs.onlinelibrary.wiley.com\/doi\/pdf\/10.1111\/febs.70277","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/febs.onlinelibrary.wiley.com\/doi\/full-xml\/10.1111\/febs.70277","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/febs.onlinelibrary.wiley.com\/doi\/pdf\/10.1111\/febs.70277","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,2,4]],"date-time":"2026-02-04T17:48:33Z","timestamp":1770227313000},"score":1,"resource":{"primary":{"URL":"https:\/\/febs.onlinelibrary.wiley.com\/doi\/10.1111\/febs.70277"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,10,5]]},"references-count":35,"journal-issue":{"issue":"3","published-print":{"date-parts":[[2026,2]]}},"alternative-id":["10.1111\/febs.70277"],"URL":"https:\/\/doi.org\/10.1111\/febs.70277","archive":["Portico"],"relation":{},"ISSN":["1742-464X","1742-4658"],"issn-type":[{"value":"1742-464X","type":"print"},{"value":"1742-4658","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,10,5]]},"assertion":[{"value":"2025-04-21","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2025-09-22","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2025-10-05","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}]}}