Fronticelli C, O'Donnell J K, Brinigar W S
Department of Biochemistry, University of Maryland, Medical School, Baltimore 21201.
J Protein Chem. 1991 Oct;10(5):495-501. doi: 10.1007/BF01025477.
A plasmid analogous to the one described by Nagai and Thogersen (Nature, 309, 810-812, 1984) has been constructed for the expression of globins in E. coli. Induction with nalidixic acid produces high yields of a fusion protein, NS1-FX-beta-globin, where NS1 represents 81 residues of a flu virus protein and FX represents a blood-clotting Factor Xa recognition sequence, Ile-Glu-Gly-Arg. This fusion protein is readily solubilized in 50 mM NaOH and remains in solution when the pH is adjusted to 8.6. Under these conditions, the fusion protein is hydrolyzed by activated Factor X, giving authentic beta-globin which can be folded in the presence of cyanohemin and native alpha-chains to produce a tetrameric hemoglobin with the functional properties of natural human hemoglobin.
已构建出一种与Nagai和Thogersen（《自然》，第309卷，810 - 812页，1984年）所描述的质粒类似的质粒，用于在大肠杆菌中表达珠蛋白。用萘啶酸诱导可产生高产量的融合蛋白NS1 - FX - β - 珠蛋白，其中NS1代表流感病毒蛋白的81个残基，FX代表凝血因子Xa识别序列Ile - Glu - Gly - Arg。这种融合蛋白很容易溶解在50 mM氢氧化钠中，当pH值调至8.6时仍保持在溶液中。在这些条件下，融合蛋白被活化的因子X水解，产生真正的β - 珠蛋白，其可在氰高铁血红素和天然α链存在的情况下折叠，以产生具有天然人血红蛋白功能特性的四聚体血红蛋白。
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