Identification of the amino acid residue of CYP27B1 responsible for binding of 25-hydroxyvitamin D3 whose mutation causes vitamin D-dependent rickets type 1.

Abstract	We previously reported the three-dimensional structure of human CYP27B1 (25-hydroxyvitamin D3 1alpha-hydroxylase) constructed by homology modeling. Using the three-dimensional model we studied the docking of the substrate, 25-hydroxyvitamin D3, into the substrate binding pocket of CYP27B1. In this study, we focused on the amino acid residues whose point mutations cause vitamin D-dependent rickets type 1, especially unconserved residues among mitochondrial CYPs such as Gln65 and Thr409. Recently, we successfully overexpressed mouse CYP27B1 by using a GroEL/ES co-expression system. In a mutation study of mouse CYP27B1 that included spectroscopic analysis, we concluded that in a 1alpha-hydroxylation process, Ser408 of mouse CYP27B1 corresponding to Thr409 of human CYP27B1 forms a hydrogen bond with the 25-hydroxyl group of 25-hydroxyvitamin D3. This is the first report that shows a critical amino acid residue recognizing the 25-hydroxyl group of the vitamin D3.
Authors	Keiko Yamamoto, Eriko Uchida, Naoko Urushino, Toshiyuki Sakaki, Norio Kagawa, Natsumi Sawada, Masaki Kamakura, Shigeaki Kato, Kuniyo Inouye, Sachiko Yamada
Journal	The Journal of biological chemistry (J Biol Chem) Vol. 280 Issue 34 Pg. 30511-6 (Aug 26 2005) ISSN: 0021-9258 [Print] United States
PMID	15972816 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References	Oligonucleotides Recombinant Proteins Glutamine Vitamin D Cholecalciferol Threonine 25-Hydroxyvitamin D3 1-alpha-Hydroxylase Calcifediol
Topics	25-Hydroxyvitamin D3 1-alpha-Hydroxylase (chemistry) Amino Acid Sequence Animals Blotting, Western Calcifediol (metabolism) Cholecalciferol (chemistry) Escherichia coli (metabolism) Glutamine (chemistry) Humans Hydrogen Bonding Kinetics Mice Models, Chemical Models, Molecular Molecular Sequence Data Mutation Oligonucleotides (chemistry) Plasmids (metabolism) Point Mutation Polymerase Chain Reaction Protein Binding Protein Conformation Protein Folding Protein Structure, Tertiary Recombinant Proteins (chemistry) Rickets (genetics) Sequence Homology, Amino Acid Spectrophotometry Substrate Specificity Threonine (chemistry) Time Factors Vitamin D (metabolism)

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