Chromatofocusing in the purification and separation of apo- and holo-(vitamin D-binding protein).
Chromatofocusing in the purification and separation of apo- and holo-(vitamin D-binding protein).
复制标题
Apo- 和 Holo-（维生素 D 结合蛋白）的纯化和分离中的色谱聚焦。
作者：
M. Keenan;R. Holmes
采用色谱聚焦法从猪血浆中纯化维生素d结合蛋白（DBP），首先采用deae -纤维素层析，然后采用DEAE-Sephadex层析，最后采用色谱聚焦法纯化。该蛋白的纯度是其血浆浓度的184倍。当血浆用[3H]钙二醇浓度的示踪剂标记时，很明显，holo- dbp和apo-DBP在色谱聚焦上没有共色谱。用纯化的载DBP与示踪剂或饱和浓度的钙二醇混合，验证了这两种形式的DBP在色谱聚焦上的分离。这种分离与它们在等电点上观察到的差异是一致的。分离载脂蛋白和全息型DBP的能力应该允许研究它们与其他结合蛋白的特定相互作用，并有助于确定这些相互作用的生理相关性。
Chromatofocusing was used to purify the vitamin D-binding protein (DBP) from pig plasma in a procedure that consisted of an initial DEAE-cellulose chromatography followed by DEAE-Sephadex chromatography, with final purification by chromatofocusing. The protein was purified 184-fold over its concentration in plasma. When the plasma was labelled with a tracer concentration of [3H]calcidiol, it was apparent that holo- and apo-DBP did not co-chromatograph on chromatofocusing. The separation of these two forms of DBP on chromatofocusing was verified by using purified apo-DBP mixed with either a tracer or a saturating concentration of calcidiol. This separation was consistent with differences observed in their isoelectric points. The ability to separate apo and holo forms of DBP should permit the study of their specific interactions with other binding proteins and help determine the physiological relevance of these interactions.