Purification and partial characterization of a nonprimate growth hormone receptor.
Purification and partial characterization of a nonprimate growth hormone receptor.
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非灵长类生长激素受体的纯化和部分表征。
DOI：
10.1016/s0021-9258(18)50441-5
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发表时间：
1979
期刊：
影响因子：
--
通讯作者：
H. Friesen
中科院分区：
文献类型：
--
作者：
M. Waters;H. Friesen
置换分析显示，孕兔肝膜具有两种类型的受体，生长激素（GH）受体结合牛生长激素的亲和常数（KA）为3 × 10（9）M-1，结合绵羊催乳素的KA为3 × 10（8）M-1，催乳素（Prl）特异性受体结合绵羊催乳素的KA为5 × 10（9）M-1。当用Triton溶解时，催乳素特异性受体的KA增加4倍，而溶解后生长激素受体的KA略微降低至2 x 10（9）M-1。亲和力的10倍差异，其结果已被利用，以促进这两种受体的分离，通过差分亲和色谱对人生长激素（hGH）的亲和凝胶。用4 M尿素从凝胶中洗脱生长激素受体，同时需要5 M MgCl 2来固定催乳素受体。优化了亲和层析条件，通过制备等电聚焦和Sepharose 6 B凝胶过滤进一步纯化GH受体，使受体纯化倍数达到8000倍以上。该物质的斯托克斯半径为62 A，与300，000的分子量一致，并且在十二烷基硫酸钠（SDS）聚丙烯酰胺凝胶上产生一个主带（75，000至80，000）和两个次带，这可以解释为指示四聚体受体。GH受体被证明是一种唾液酸糖蛋白（或与唾液酸糖蛋白密切相关），通过分析等电点为4.6。使用高度纯化受体的特异性研究证实了最初的假设，即与催乳素特异性受体相反，该受体能够以高亲和力结合牛生长激素（bGH），以低亲和力结合绵羊催乳素（oPrl）。
Pregnant rabbit liver membranes have been shown to possess two types of receptors by displacement analysis, a growth hormone (GH) receptor which binds bovine growth hormone with an affinity constant (KA) of 3 x 10(9) M-1 and ovine prolactin with a KA of 3 x 10(8) M-1, and a prolactin (Prl)-specific receptor which binds ovine prolactin with a KA of 5 x 10(9) M-1. The prolactin-specific receptor when solubilized with Triton exhibits a 4-fold increase in the its KA while the KA of the growth hormone receptor decreases slightly to 2 x 10(9) M-1 after solubilization. The 10-fold difference in affinity which results has been exploited to facilitate the separation of these two receptors by differential affinity chromatography on human growth hormone (hGH) affinity gels. The growth hormone receptor is eluted from the gel with 4 M urea while 5 M MgCl2 is required to elute the prolactin receptor. Conditions of affinity chromatography have been optimized, and further purification of the GH receptor by preparative isoelectric focusing and Sepharose 6B gel filtration resulted in a more than 8000-fold purification of the receptor. This material had a Stokes radius of 62 A, consistent with a molecular weight of 300,000 and gave one main band (75,000 to 80,000) and two minor bands on sodium dodecyl sulfate (SDS) polyacrylamide gels, which could be interpreted as indicating a tetrameric receptor. The GH receptor was shown to be a sialoglycoprotein (or closely associated with sialoglycoprotein) by analytical isoelectric focusing with an isoelectric point of 4.6. Specificity studies with the highly purified receptor confirmed the initial hypothesis that this receptor is capable of binding bovine growth hormone (bGH) with high affinity and ovine prolactin (oPrl) with low affinity, in contrast to the prolactin-specific receptor.