ESPE Abstracts

Background: CYP27B1 converts 25-hydroxyvitamin D₃ to active 1,25-dihydroxyvitamin D₃, playing a vital role in calcium homeostasis and bone growth. Vitamin D-dependent rickets type 1 (VDDR-1) is a rare autosomal recessive disorder caused by mutations in CYP27B1.

Objective and hypotheses: Enzymatic and structural analysis of mutations in a patient with calcipenic rickets.

Method: Two siblings presented with calcipenic rickets and inappropriately normal 1.25-dihydroxyvitamin D₃ levels. CYP27B1 gene analysis showed compound heterozygous mutations confirming VDDR-1. We studied wild type CYP27B1 and mutations H441Y and R459L by computational homology modeling, molecular dynamics simulations and functional studies using a luciferase assay. The patient was successfully treated with calcitriol.

Results: Mitochondrial cytochrome P450s require adrenodoxin (FDX1) and adrenodoxin reductase. We created models of CYP27B1-FDX1 complex which revealed negative effects of mutations H441Y and R459L. Upon structural analysis, near-identical folds, protein contact areas, and orientations of heme/iron-sulfur cluster suggested that both mutations destabilize the CYP27B1-FDX1 complex by negating directional interactions with adrenodoxin. This system is highly sensitive to small local changes modulating the binding/unbinding of adrenodoxin, and electron-transporting efficiency changes with mutations at the surface. Functional assays confirmed this hypothesis and showed severe loss of activity of CYP27B1 by both mutations.

Conclusion: This is the first report of mutations in CYP27B1 causing VDDR-1 by affecting protein-protein interactions with FDX1 that results in reduced CYP27B1 activities. Detailed characterization of mutations in CYP27B1 is required for understanding the novel molecular mechanisms causing VDDR-1.