Gonzalez-Gronow M, Robbins K C
Biochemistry. 1984 Jan 17;23(2):190-6. doi: 10.1021/bi00297a003.
mRNA was isolated from total RNA of monkey liver by oligo(dT)-cellulose chromatography and was translated in a rabbit reticulocyte cell-free system. Analysis of the translation products immunoprecipitated with specific antibodies to monkey plasma plasminogen revealed a molecule with characteristics similar to those of native plasminogen. The purification of the mRNA by centrifugation on sucrose gradients indicated the presence of plasminogen mRNAs in both the 23S and 18S RNA fractions. Both plasminogen mRNAs can be further purified by chromatography on Sepharose 4B. Affinity chromatography of the proteins synthesized in vitro by total mRNA from liver, as well as by the purified mRNAs, on L-lysine-substituted Sepharose revealed that both major plasma plasminogen forms (1 and 2) are synthesized, as precursors, in the system. The in vitro synthesized plasminogen is similar in its physical and chemical properties to native plasma plasminogen as determined by its ability to bind to L-lysine-substituted Sepharose and its molecular interaction with streptokinase. The purified mRNAs were also translated in the presence of dog pancreas microsomal membranes, and and fractionated on concanavalin A-Sepharose. The 23S mRNA directed the synthesis of a plasminogen molecule similar to the circulating plasma plasminogen form 1, whereas the 18S mRNA directed the synthesis of a molecule similar to the circulating plasma plasminogen form 2. Our evidence indicates that the synthesis of the two major circulating plasma plasminogen forms is directed in the liver by separate mRNAs.
通过寡聚（dT）-纤维素层析从猴肝总RNA中分离出mRNA，并在兔网织红细胞无细胞系统中进行翻译。用针对猴血浆纤溶酶原的特异性抗体免疫沉淀翻译产物进行分析，发现了一种具有与天然纤溶酶原相似特征的分子。通过在蔗糖梯度上离心纯化mRNA，结果表明在23S和18S RNA组分中均存在纤溶酶原mRNA。两种纤溶酶原mRNA均可通过琼脂糖凝胶4B层析进一步纯化。用肝总mRNA以及纯化后的mRNA体外合成的蛋白质在L-赖氨酸取代的琼脂糖凝胶上进行亲和层析，结果显示该系统中合成了两种主要的血浆纤溶酶原形式（1和2）的前体。通过其与L-赖氨酸取代的琼脂糖凝胶结合的能力及其与链激酶的分子相互作用测定，体外合成的纤溶酶原在物理和化学性质上与天然血浆纤溶酶原相似。纯化后的mRNA也在犬胰腺微粒体膜存在的情况下进行翻译，并在伴刀豆球蛋白A-琼脂糖凝胶上进行分级分离。23S mRNA指导合成一种与循环血浆纤溶酶原形式1相似的纤溶酶原分子，而18S mRNA指导合成一种与循环血浆纤溶酶原形式2相似的分子。我们的证据表明，肝脏中两种主要的循环血浆纤溶酶原形式的合成是由不同的mRNA指导的。
