Ferguson B, Pritchard M L, Feild J, Rieman D, Greig R G, Poste G, Rosenberg M
J Biol Chem. 1985 Mar 25;260(6):3652-7.
A segment of the coding sequence of the Abelson murine leukemia virus transforming gene (v-abl) has been inserted into a plasmid vector that allows its efficient and regulated expression in Escherichia coli. The product of the v-abl-derived coding sequence, designated p60v-abl, accumulated to a level of approximately 10% of total E. coli protein. A procedure is described for the isolation of p60v-abl from E. coli that yields about 50 micrograms of p60v-abl/g wet weight of E. coli. p60v-abl was capable of autophosphorylation and phosphorylating certain E. coli proteins specifically at tyrosine residues. The E. coli-expressed p60v-abl specifically phosphorylated tyrosine residues on casein and angiotensin II. The Km and Vmax values for ATP, casein, and angiotensin II in the p60v-abl kinase reaction have been determined and compared to values reported for other tyrosine-specific kinases. The expression system and isolation procedure described here permit the preparation of functional p60v-abl in quantities sufficient for detailed physical and biochemical characterization and examination of its biological action(s).
阿贝尔逊鼠白血病病毒转化基因（v-abl）的一段编码序列已被插入到一个质粒载体中，该载体能使其在大肠杆菌中高效且受调控地表达。v-abl衍生编码序列的产物，命名为p60v-abl，其积累量达到大肠杆菌总蛋白的约10%。本文描述了一种从大肠杆菌中分离p60v-abl的方法，每克湿重的大肠杆菌可产生约50微克的p60v-abl。p60v-abl能够进行自身磷酸化，并特异性地在酪氨酸残基上磷酸化某些大肠杆菌蛋白。大肠杆菌表达的p60v-abl能特异性地磷酸化酪蛋白和血管紧张素II上的酪氨酸残基。已测定了p60v-abl激酶反应中ATP、酪蛋白和血管紧张素II的米氏常数（Km）和最大反应速度（Vmax）值，并与其他酪氨酸特异性激酶报道的值进行了比较。本文所述的表达系统和分离方法能够制备出足够量的功能性p60v-abl，用于详细的物理和生化特性分析以及对其生物学作用的研究。
