Watson D H
Biochem J. 1980 Feb 1;185(2):463-71. doi: 10.1042/bj1850463.
Coupled transcription and translation of plasmid-ColE1 DNA in vitro under optimized conditions gave one major product. This has an apparent weight of 71 000, the same N-terminal sequence as colicin E1 and was not digested by deoxyribonuclease or ribonuclease. It differed from colicin E1 in its C-terminal residue and amino acid composition. It had lower specific activities in cell killing and in the fluorescence-enhancement in vitro assay of Phillips & Cramer [(1973) Biochemistry 12, 1170--1176] than did colicin E1, but both proteins bound in equimolar amounts to colicin-sensitive and colicin-resistant cells. The product of plasmid-ColE1-DNA-directed protein synthesis was converted into a protein indistinguishable in structure and activity from colicin E1 by incubation in the reaction mixture, after deoxyribonuclease and ribonuclease treatment, for a further 20 h at 37 degrees C. A protein with similar properties to the 71 000-dalton product in vitro was identified in extracts of a ColE1+ colicin-tolerant mutant of Escherichia coli K12. It is concluded that this protein probably represents a pre-form of colicin E1 which may be involved in colicin-E1 secretion or cellular colicin-E1 immunity in colicin-E-producing cells, or both of these processes.
在优化条件下对质粒 - ColE1 DNA进行体外转录和翻译偶联反应，产生了一种主要产物。该产物的表观分子量为71000，与大肠杆菌素E1具有相同的N端序列，且不被脱氧核糖核酸酶或核糖核酸酶消化。它在C端残基和氨基酸组成上与大肠杆菌素E1不同。在细胞杀伤以及菲利普斯和克莱默（1973年，《生物化学》12卷，1170 - 1176页）的体外荧光增强测定中，其比活性低于大肠杆菌素E1，但这两种蛋白质与大肠杆菌素敏感细胞和抗大肠杆菌素细胞的结合量均为等摩尔。在脱氧核糖核酸酶和核糖核酸酶处理后的反应混合物中于37℃再孵育20小时后，质粒 - ColE1 - DNA指导的蛋白质合成产物转化为一种在结构和活性上与大肠杆菌素E1无法区分的蛋白质。在大肠杆菌K12的ColE1 + 大肠杆菌素耐受突变体提取物中鉴定出一种在体外具有与71000道尔顿产物相似性质的蛋白质。得出的结论是，这种蛋白质可能代表大肠杆菌素E1的一种前体形式，它可能参与大肠杆菌素E产生细胞中的大肠杆菌素E1分泌或细胞对大肠杆菌素E1的免疫，或者这两个过程。
