期刊
BIOPHYSICAL JOURNAL
卷 90, 期 10, 页码 3389-3409出版社
CELL PRESS
DOI: 10.1529/biophysj.105.069369
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Human CYP27A1 is a mitochondrial cytochrome P450, which is principally found in the liver and plays important roles in the biological activation of vitamin D-3 and in the biosynthesis of bile acids. We have applied a systematic analysis of hydrogen bonding patterns in 11 prokaryotic and mammalian CYP crystal structures to construct a homology-based model of CYP27A1. Docking of vitamin D-3 structures into the active site of this model identified potential substrate contact residues in the F-helix, the beta-3 sheet, and the beta-5 sheet. Site-directed mutagenesis and expression in COS-1 cells confirmed that these positions affect enzymatic activity, in some cases shifting metabolism of 1 alpha-hydroxyvitamin D-3 to favor 25- or 27-hydroxylation. The results suggest that conserved hydrophobic residues in the beta-5 hairpin help define the shape of the substrate binding cavity and that this structure interacts with Phe-248 in the F-helix. Mutations directed toward the beta-3a strand suggested a possible heme-binding interaction centered on Asn-403 and a structural role for substrate contact residues Thr-402 and Ser-404.
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