Production and purification of recombinant human inhibin and activin
Production and purification of recombinant human inhibin and activin
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DOI：
10.1677/joe.0.1720199
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发表时间：
2002-01-01
影响因子：
4
通讯作者：
Woodruff, TK
中科院分区：
文献类型：
--
作者：
Pangas, SA;Woodruff, TK
抑制素和激活素是具有多种生理功能的蛋白质激素，包括调节垂体FSH的分泌，与转化生长因子-P基因家族的其他成员一样，它们经历较大的前体分子的加工以及组装成功能性二聚体。天然来源的抑制素和激活素的分离只能产生有限数量的生物活性蛋白。为了大量纯化重组人抑制素和激活素，我们在自备的生物反应器中利用稳定表达的细胞系来生产蛋白质。这些细胞每天产生大约200微克/毫升的重组人抑制素。用柱层析法纯化条件细胞培养液，可获得成熟的32-34 kDa抑制素A和28 kDa激活素A二聚体。纯化的重组蛋白保持了传统体外测定的生物活性，包括对大鼠垂体前叶培养物中FSH的调节和对组织培养细胞中激活素反应启动子p3TP-Luc启动子活性的调节。这些蛋白质将对未来抑制素和激活素功能的分析有价值，并已分发给美国国家激素和多肽计划。
Inhibin and activin are protein hormones with diverse physiological roles including the regulation of pituitary FSH secretion, Like other members of the transforming growth factor-P gene family, they undergo processing from larger precursor molecules as well as assembly into functional dimers. Isolation of inhibin and activin front natural sources can only produce limited quantities of bioactive protein. To purify large-scale quantities of recombinant human inhibin and activin, we have utilized stably transfected cell lines in self-contained bioreactors to produce protein. These cells produce approximately 200 mug/ml per day total recombinant human inhibin. Conditioned cell media can be purified through column chromatography resulting in dimeric mature 32-34 kDa inhibin A and 28 kDa activin A. The purified recombinant proteins maintain their biological activity as measured by traditional in vitro assays including the regulation of FSH in rat anterior pituitary cultures and the regulation of promoter activity of the activin-responsive promoter p3TP-luc in tissue culture cells. These proteins will be valuable for future analysis of inhibin and activin function and have been distributed to the US National Hormone and Peptide Program.