Zettlmeissl G, Conradt H S, Nimtz M, Karges H E
Research Laboratories of Behringwerke AG, Marburg, Federal Republic of Germany.
J Biol Chem. 1989 Dec 15;264(35):21153-9.
Biochemical and physiochemical properties of recombinant human antithrombin III were examined. This protein, produced in Chinese hamster ovary cells, showed a conformation apparently identical with the natural product isolated from human plasma when examined by circular dichroism, UV absorbance, and fluorescence spectroscopy. Comparison of the NH2-terminal sequences of recombinant and human plasma-derived antithrombin III showed that on synthesis and secretion of the recombinant protein from Chinese hamster ovary cells the signal peptide is correctly cleaved by the corresponding endoplasmic signal peptidase. The recombinant antithrombin III has identical properties in heparin binding and biological activities as determined in vitro by two-dimensional immunoelectrophoresis, progressive inhibitor, and heparin cofactor assays. Analysis of the carbohydrate portion of recombinant antithrombin III synthesized in Chinese hamster ovary cells revealed glycosylation of the complex type. Characterization of the oligosaccharide chains present in the recombinant protein reveals three major fractions, A (20%), B (60%), and C (20%). Fraction A contains tri- and tetraantennary complex-type oligosaccharides, fraction B contains biantennary oligosaccharides, and fraction C partially truncated biantennary structures. Pharmacokinetic studies with recombinant and plasma-derived antithrombin III in rabbits showed that the clearance behavior of both proteins is very similar and can be described by a double exponential decrease with almost identical kinetic parameters.
对重组人抗凝血酶III的生化和物理化学性质进行了研究。这种在中国仓鼠卵巢细胞中产生的蛋白质，通过圆二色性、紫外吸收和荧光光谱检测时，其构象与从人血浆中分离出的天然产物明显相同。重组抗凝血酶III与人血浆来源的抗凝血酶III的NH2末端序列比较表明，当重组蛋白从中国仓鼠卵巢细胞合成并分泌时，信号肽被相应的内质网信号肽酶正确切割。通过二维免疫电泳、渐进抑制剂和肝素辅因子测定法在体外测定，重组抗凝血酶III在肝素结合和生物活性方面具有相同的性质。对在中国仓鼠卵巢细胞中合成的重组抗凝血酶III的碳水化合物部分进行分析，发现其为复合型糖基化。对重组蛋白中存在的寡糖链进行表征，发现有三个主要部分，A（20%）、B（60%）和C（20%）。A部分含有三触角和四触角复合型寡糖，B部分含有双触角寡糖，C部分含有部分截短的双触角结构。用重组抗凝血酶III和血浆来源的抗凝血酶III对兔子进行的药代动力学研究表明，两种蛋白质的清除行为非常相似，可用双指数下降来描述，动力学参数几乎相同。
