scholarly journals Electrospray mass spectrometry for the direct accurate mass measurement of ligands in complex with the retinoid X receptor α ligand binding domain

2005 ◽  
Vol 16 (10) ◽  
pp. 1631-1640 ◽  
Author(s):  
Johan Lengqvist ◽  
Gunvor Alvélius ◽  
Hans Jörnvall ◽  
Jan Sjövall ◽  
Thomas Perlmann ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ligand Binding ◽  
Electrospray Mass Spectrometry ◽  
Mass Measurement ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Retinoid X Receptor ◽  
Accurate Mass Measurement ◽  
Accurate Mass

scholarly journals Hereditary 1,25-Dihydroxyvitamin D Resistant Rickets due to a Mutation Causing Multiple Defects in Vitamin D Receptor Function

Endocrinology ◽  
2004 ◽  
Vol 145 (11) ◽  
pp. 5106-5114 ◽  
Author(s):  
Peter J. Malloy ◽  
Rong Xu ◽  
Lihong Peng ◽  
Sara Peleg ◽  
Abdullah Al-Ashwal ◽  
...  
Keyword(s):  
Vitamin D ◽  
Ligand Binding ◽  
Vitamin D Receptor ◽  
Novel Mutation ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Retinoid X Receptor ◽  
Vdr Gene ◽  
Dihydroxyvitamin D3 ◽  
Resistant Rickets

Abstract Hereditary vitamin D-resistant rickets (HVDRR) is an autosomal recessive disease caused by mutations in the vitamin D receptor (VDR). We studied a young Saudi Arabian girl who exhibited the typical clinical features of HVDRR, but without alopecia. Analysis of her VDR gene revealed a homozygous T to C mutation in exon 7 that changed isoleucine to threonine at amino acid 268 (I268T). From crystallographic studies of the VDR ligand-binding domain, I268 directly interacts with 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and is involved in the hydrophobic stabilization of helix H12. We recreated the I268T mutation and analyzed its effects on VDR function. In ligand binding assays, the I268T mutant VDR exhibited an approximately 5- to 10-fold lower affinity for [3H]1,25(OH)2D3 compared with the wild-type (WT) VDR. The I268T mutant required approximately a 65-fold higher concentration of 1,25(OH)2D3 to be equipotent in gene transactivation. Both retinoid X receptor heterodimerization and coactivator binding were reduced in the I268T mutant. Analogs of 1,25(OH)2D3 have been proposed as potential therapeutics for patients with HVDRR. Interestingly, in protease sensitivity assays, treatment with the potent vitamin D analog, 20-epi-1,25(OH)2D3, stabilized I268T mutant proteolytic fragments better than 1,25(OH)2D3. Moreover, 20-epi-1,25(OH)2D3 restored transactivation of the I268T mutant to levels exhibited by WT VDR treated with 1,25(OH)2D3. In conclusion, we describe a novel mutation, I268T, in the VDR ligand-binding domain that alters ligand binding, retinoid X receptor heterodimerization, and coactivator binding. These combined defects in VDR function cause resistance to 1,25(OH)2D3 action and result in the syndrome of HVDRR.

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