Purification by affinity chromatography and immunological characterization of a 110kDa component of the chick oviduct progesterone receptor.
Purification by affinity chromatography and immunological characterization of a 110kDa component of the chick oviduct progesterone receptor.
复制标题
通过亲和色谱法纯化鸡输卵管黄体酮受体的 110kDa 成分并进行免疫学表征。
作者：
J. Renoir;J. Mester;T. Buchou;M. Catelli;P. Tuohimaa;N. Binart;I. Joab;C. Radanyi;E. Baulieu
鸡输卵管孕酮受体（PR）的110 kDa组分已根据电泳标准和比活性（假设一个孕酮结合位点/110 kDa）纯化至均一。该程序涉及0.3 M-KCl制备的胞质溶胶的亲和层析，然后进行DEAE-Sephacel层析（在0.2 M-KCl下洗脱）。就结合活性而言，最终产率为约12%。110 kDa组分的性质表明，它与先前描述的“B”亚基相同[斯托克斯半径约6.1 nm;沉降系数，（S20，w）约4S;摩擦比约1.77]。它与IgG-G3多克隆抗体反应，但不与针对8 S精氨酸稳定的鸡输卵管PR的BF 4单克隆抗体反应，并与其90 kDa组分反应。另一种与“A”亚基相对应的甾醇结合组分，也如前所述，在约0.08 M-KCl下从DEAE-Sephacel柱上洗脱下来，并含有分子量约为0.08 M-KCl的肽。75- 80 kDa，具有S20，在蔗糖梯度中w约4S。该组分也被IgG-G3识别，但不被BF 4识别;它在酶结合活性方面非常不稳定。
A 110kDa component of the chick oviduct progesterone receptor (PR) has been purified to homogeneity according to electrophoretic criteria and specific activity (assuming one progestagen-binding site/110kDa). The procedure involved affinity chromatography of 0.3 M-KCl-prepared cytosol, followed by DEAE-Sephacel chromatography (elution at 0.2 M-KCl). The final yield was about 12% in terms of binding activity. Properties of the 110kDa component indicate that it is identical with the 'B' subunit described previously [Stokes radius approximately 6.1 nm; sedimentation coefficient, (S20, w) approximately 4S; frictional ratio approximately 1.77]. It reacted with the IgG-G3 polyclonal antibody, but not with BF4 monoclonal antibody raised against the 8S molybdate-stabilized chick oviduct PR and reacting with its 90kDa component. Another progesterone-binding component, corresponding to the 'A' subunit, also previously described, was eluted from the DEAE-Sephacel column at approximately 0.08 M-KCl, and contained a peptide of molecular mass approx. 75-80kDa, which had S20, w approximately 4S in a sucrose gradient. This component was also recognized by IgG-G3, but not by BF4; it was very unstable in terms of hormone-binding activity.