25-Hydroxy[26,27-methyl-3H]vitamin D3-3 beta-(1,2-epoxypropyl)ether: an affinity labeling reagent for human vitamin D-binding protein.
25-Hydroxy[26,27-methyl-3H]vitamin D3-3 beta-(1,2-epoxypropyl)ether: an affinity labeling reagent for human vitamin D-binding protein.
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25-羟基[26,27-甲基-3H]维生素 D3-3 β-(1,2-环氧丙基)醚：人维生素 D 结合蛋白的亲和标记试剂。
DOI：
10.1006/abbi.1995.1323
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Ray,R
中科院分区：
文献类型：
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作者：
Swamy,N;Ray,R
维生素d结合蛋白（Vitamin d binding protein， DBP）主要参与维生素d3及其各种代谢物与目标器官和组织的结合和运输。这表现在舒张肽与维生素d3及其代谢物高亲和力结合的能力上。本研究开发了25-羟基维生素D3-3β-（1,2-环氧丙基）醚（25- oh - d3 -环氧化物）作为人舒张压（hDBP）的亲和标记试剂。25-OH-D3-环氧化物与hDBP的竞争性放射配体结合实验表明，这种类似物的结合亲和力与25-羟基维生素D3（25-OH-D3）相似。25-羟基[26(27)-3H]维生素D3-3β-（1,2-环氧丙基）醚[[3H]25- oh - d3 -环氧化物]与hDBP共价标记的蛋白质孵育。当在25-OH-D3大量过量的情况下进行孵育时，完全去除标记。当含hDBP的人Cohn IV部分与[3H]25- oh - d3 -环氧化物孵育时，标记单个蛋白带，对应于]hDBP。在大量25-OH-D3的存在下，标记被完全抹掉。然而，等量的7-脱氢胆固醇对标签没有影响。这些结果表明[3H]25- oh - d3 -环氧化物很可能标记了hDBP的维生素D甾醇结合域。
Vitamin D-binding protein (DBP) is primarily involved in the binding and transportation of vitamin D3and its various metabolites to target organs and tissues. This is manifested by the ability of DBP to bind vitamin D3and its metabolites with high affinity. In the present study we developed 25-hydroxyvitamin D3-3β-(1,2-epoxypropyl)ether (25-OH-D3-epoxide) as an affinity labeling reagent of human DBP (hDBP). Competitive radioligand binding assays of 25-OH-D3-epoxide with hDBP demonstrated that the binding affinity of this analog was similar to that of 25-hydroxyvitamin D3(25-OH-D3). Incubation of 25-hydroxy[26(27)-3H]vitamin D3-3β-(1,2-epoxypropyl)ether [[3H]25-OH-D3-epoxide] with hDBP covalently labeled the protein. When the incubation was carried out in the presence of a large excess of 25-OH-D3, labeling was removed completely. When human Cohn IV fraction, containing hDBP, was incubated with [3H]25-OH-D3-epoxide a single protein band, corresponding to]hDBP, was labeled. Labeling was completely obliterated in the presence of a large amount of 25-OH-D3. However, an equivalent amount of 7-dehydrocholesterol had no effect on labeling. These results demonstrated that [3H]25-OH-D3-epoxide most probably labeled the vitamin D sterol-binding domain of hDBP.