Purification of plasminogen activator from Rous sarcoma virus transformed chick embryo fibroblasts treated with the tumor promoter phorbol 12-myristate 13-acetate.
Purification of plasminogen activator from Rous sarcoma virus transformed chick embryo fibroblasts treated with the tumor promoter phorbol 12-myristate 13-acetate.
复制标题
从用肿瘤启动子佛波醇 12-肉豆蔻酸酯 13-乙酸酯处理的劳斯肉瘤病毒转化的鸡胚成纤维细胞中纯化纤溶酶原激活剂。
DOI：
10.1021/bi00565a001
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发表时间：
1980
期刊：
影响因子：
2.9
通讯作者：
J. Quigley
中科院分区：
文献类型：
--
作者：
R. Goldfarb;J. Quigley
罗纳德H.劳氏肉瘤病毒（RSV）转化的鸡胚成纤维细胞产生一种特异的丝氨酸蛋白酶，纤溶酶原激活剂，其水平比未转化的培养物高25-100倍。用肿瘤促进剂佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）处理转化的培养物进一步增强了转化的培养物中纤溶酶原激活剂的产生，从而允许收获足够量的粗酶用于纯化研究[Goldfarb，R. H、& Quigley，JP（1978）CancerRes. 38，4601]，从PMA处理的转化培养物的培养上清液中收获的纤溶酶原激活剂现已通过对氨基苯甲脒-Sepharose 4 B上的多重亲和层析、Sephadex G-100凝胶过滤和磺丙基阳离子交换层析纯化至表观均一性。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分析表明，纤溶酶原激活剂被分离为两种酶活性形式。溶血酶具有刺激细胞生长、增强细胞迁移、修饰细胞内和细胞外蛋白质组分的能力，是直接或间接负责恶性细胞的几种表型特征的催化调节分子的候选者。蛋白水解活性水平的增加通常与体内恶性肿瘤和培养物中的恶性转化相关[参见Quigley（1979）的综述]。此外，许多研究表明，当用外源性蛋白水解酶处理非转化细胞时，会暂时显示出转化细胞的几种表型特征[见Noonan的综述（1978）]。相反，几种特异性蛋白酶抑制剂，在加入培养物中的转化细胞后，
Ronald H. Goldfarb* andJames P. Quigley* abstract: Rous sarcoma virus (RSV) transformed chick embryo fibroblasts produce a specific serine protease, plas-minogen activator at levels 25-100-fold greater than non-transformed cultures. Treatment of transformed cultures with the tumor promoter phorbol 12-myristate 13-acetate (PMA) further enhances the production of plasminogen activator in the transformed cultures thereby permitting sufficient quan-tities of crude enzyme to be harvested for purification studies [Goldfarb, R. H., & Quigley, JP (1978) CancerRes. 38, 4601], The plasminogen activator harvested from the culture supernatants of PMA-treated, transformedcultures has now been purified to apparent homogeneity by multiple affinity chromatography on p-aminobenzamidine-Sepharose 4B, Sephadex G-100 gel filtration, and sulfopropyl cation-exchange chromatography. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicates that plasminogen activator is isolated as two enzymatically active forms. TheI^ oteolytic enzymes, which have the ability to stimulate cell growth, enhance cellular migration, and modify cellular and extracellular protein components, are candidates as catalytic regulatory molecules that could be responsible, directly or indirectly, for several of the phenotypic features of the ma-lignant cell. Increased levels of proteolytic activity are often associated with malignanttumors in vivo and malignant transformation in culture [see review by Quigley (1979)]. In addition, numerous studies have shown that nontransformed cells, when treated with exogenously added proteolytic en-zymes, transiently display several phenotypic propertiesthat are characteristic of transformed cells [see review by Noonan (1978)]. Conversely, several specific protease inhibitors, upon addition to transformed cells in culture, partially restore