Bouillon R, Van Baelen H, Tan B K, De Moor P
J Biol Chem. 1980 Nov 25;255(22):10925-30.
The binding protein for 15-hydroxyvitamin D3 (25-OH-D3) was isolated from chick serum. The molecular weight was 60,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, without the existence of sub-units. Its sedimentation coefficient was 4.3 S. The isoelectric points of the major and minor components were 6.1 and 5.9, respectively. Addition of 25-hydroxyvitamin D3 causes an anodal shift in the isoelectric point of the two forms of the binding protein. Heating at 60 degrees C slowly destroys the binding protein but the addition of vitamin D metabolites increases its thermostability. The binding protein has probably a single binding site for all vitamin D metabolites and its association constant for 25-OH-D3 at 4 degrees C and pH 7.4 is 10(9) M-1. The same affinity is found for 24,25-dihydroxyvitamin D3 but 25-hydroxyvitamin D3 and 1,25-dihydroxyvitamin D3 have a 3- and 10-fold lower affinity, respectively. The amino acid composition is very similar to that of the human and rat binding protein. The concentration of chick 25-hydroxyvitamin D3-binding protein was higher in egg-laying hens (10 microM) than in immature hens (4 microM) or immature or adult roosters (4 microM). Chick plasma contains virtually only a 4 S form of the 25-hydroxyvitamin D3-binding protein but partially hemolyzed chick serum also contains a 6 S form. Addition of pure actin to chick plasma or serum converts all 4 S form to the 6 S form. The serum 25-hydroxyvitamin D3-binding protein is thus heterogenous since it exists in two 4 S forms (with different pI) and a 6 S form (complexed with actin). No evidence was found for the existence of a separate vitamin D3-binding protein.
从鸡血清中分离出了15-羟基维生素D3（25-OH-D3）结合蛋白。在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳上，其分子量为60,000，不存在亚基。其沉降系数为4.3 S。主要和次要成分的等电点分别为6.1和5.9。添加25-羟基维生素D3会导致两种形式的结合蛋白的等电点向阳极移动。60℃加热会缓慢破坏结合蛋白，但添加维生素D代谢物会增加其热稳定性。该结合蛋白可能对所有维生素D代谢物都有一个单一结合位点，其在4℃和pH 7.4时对25-OH-D3的缔合常数为10⁹ M⁻¹。对24,25-二羟基维生素D3也有相同的亲和力，但25-羟基维生素D3和1,25-二羟基维生素D3的亲和力分别低3倍和10倍。其氨基酸组成与人和大鼠的结合蛋白非常相似。产蛋母鸡中鸡25-羟基维生素D3结合蛋白的浓度（10微摩尔）高于未成熟母鸡（4微摩尔）或未成熟或成年公鸡（4微摩尔）。鸡血浆实际上只含有25-羟基维生素D3结合蛋白的4 S形式，但部分溶血的鸡血清也含有6 S形式。向鸡血浆或血清中添加纯肌动蛋白会将所有4 S形式转化为6 S形式。因此，血清25-羟基维生素D3结合蛋白是异质的，因为它以两种4 S形式（具有不同的pI）和一种6 S形式（与肌动蛋白复合）存在。未发现存在单独的维生素D3结合蛋白的证据。
