Buffered non-fermenter system for lab-scale production of secreted recombinant his-tagged proteins in Saccharomyces cerevisiae
Buffered non-fermenter system for lab-scale production of secreted recombinant his-tagged proteins in Saccharomyces cerevisiae
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DOI：
10.2144/02336pt02
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发表时间：
2002-12-01
期刊：
影响因子：
2.7
通讯作者：
Twining, SS
中科院分区：
文献类型：
--
作者：
Ngamkitidechakul, C;Twining, SS
利用分泌系统表达重组蛋白可以最大限度地减少污染宿主蛋白的共纯化。在酵母α因子分泌系统中生产his标记的重组蛋白以前需要一个发酵罐系统来控制酵母培养物的pH等生长条件。我们描述了一种廉价的非发酵系统，用于在酿酒酵母中生产分泌的重组his标记蛋白，该系统使用缓冲的低蛋白胨YP甘油培养基，不干扰固定化金属亲和层析。Maspin是一种肿瘤抑制丝氨酸蛋白酶，作为分泌的n端His/ FLAG(R)标记蛋白表达。可溶性活性重组蛋白的纯化只需要离心、超滤浓缩和Ni2+亲和层析。该体系的纯化蛋白产量为3-5 mg/L培养基。
Expression of recombinant proteins using a secretion system can minimize co-purification of contaminating host proteins. Production of His-tagged recombinant proteins in the yeast alpha-factor secretion system has previously required a fermenter system to control the growth conditions such as pH of the yeast culture. We describe an inexpensive non-fermenter system for the production of secreted recombinant His-tagged proteins in Saccharomyces cerevisiae that uses a buffered low peptone YP glycerol medium, which does not interfere with immobilized metal affinity chromatography. Maspin, a tumor suppressor serpin, was expressed as a secreted N-terminal His/ FLAG(R)-tagged protein. Purification of the soluble active recombinant protein only requires centrifugation, concentration by ultrafiltration, and Ni2+ affinity chromatography. Purified protein yields of this system are 3-5 mg/L culture medium.