Mason A B, Funk W D, MacGillivray R T, Woodworth R C
Department of Biochemistry, University of Vermont College of Medicine, Burlington 05405.
Protein Expr Purif. 1991 Apr-Jun;2(2-3):214-20. doi: 10.1016/1046-5928(91)90074-s.
Expression of the amino-terminal lobe of human serum transferrin secreted into the culture medium by transformed baby hamster kidney (BHK) cells has been increased from the levels reported originally of 10-15 micrograms/ml to 55-120 micrograms/ml. Use of the serum substitute, Ultraser G, has facilitated isolation of the recombinant protein, resulting in approximately 80% recovery of expressed hTF/2N from the culture medium. In the three experiments described, 300-750 mg of recombinant protein was collected over a period of 25 days from five expanded surface roller bottles each containing 200 ml of medium (seven to nine collections). The use of alginate beads to encapsulate the transformed BHK cells provided no advantage over normal culturing over 25 days. A lag in production resulting in 30% less recombinant protein over this time period was observed. The production and isolation procedures described are easily handled by one person. The system is amenable to incorporation of isotopically substituted amino acids useful in NMR studies.
经转化的幼仓鼠肾（BHK）细胞分泌到培养基中的人血清转铁蛋白氨基末端叶的表达量，已从最初报道的10 - 15微克/毫升提高到55 - 120微克/毫升。使用血清替代品Ultraser G有助于重组蛋白的分离，从培养基中回收表达的hTF/2N的回收率约为80%。在所描述的三个实验中，在25天的时间里，从五个每个装有200毫升培养基的扩展表面滚瓶（进行七至九次收集）中收集到了300 - 750毫克的重组蛋白。使用藻酸盐珠包封经转化的BHK细胞，在25天的培养过程中与正常培养相比没有优势。观察到产量出现滞后，在此期间重组蛋白产量减少了30%。所描述的生产和分离程序一个人就很容易操作。该系统适合纳入可用于核磁共振研究的同位素取代氨基酸。
