Hickson I D, Arthur H M, Bramhill D, Emmerson P T
Mol Gen Genet. 1983;190(2):265-70. doi: 10.1007/BF00330649.
We have shown that the uvrD gene product, previously identified in maxicell extracts as a 73 kilodalton protein, copurifies with single stranded DNA-dependent ATPase and ATP-dependent DNA helicase activities. This protein is specifically precipitated from maxicell extracts by antibodies raised against DNA helicase II. In order to facilitate purification of the UvrD protein we have subcloned the uvrD gene into a plasmid vector in which its transcription is under the control of the phage lambda leftward promoter. Using cells harbouring this recombinant plasmid as a source of elevated levels of the UvrD protein we have purified this protein to homogeneity by a simple, rapid procedure. The purified protein has single stranded DNA-dependent ATPase activity and ATP-dependent DNA helicase activity, and both activities are specifically inactivated by antibodies raised against DNA helicase II. We conclude that DNA helicase II is the uvrD gene product.
我们已经证明,uvrD基因产物,先前在大细胞提取物中被鉴定为一种73千道尔顿的蛋白质,与单链DNA依赖性ATP酶和ATP依赖性DNA解旋酶活性共同纯化。这种蛋白质可被针对DNA解旋酶II产生的抗体从大细胞提取物中特异性沉淀。为了便于纯化UvrD蛋白,我们已将uvrD基因亚克隆到一个质粒载体中,其转录受噬菌体λ左向启动子的控制。使用携带这种重组质粒的细胞作为UvrD蛋白高水平的来源,我们通过一个简单、快速的程序将这种蛋白纯化至同质。纯化后的蛋白具有单链DNA依赖性ATP酶活性和ATP依赖性DNA解旋酶活性,并且这两种活性都被针对DNA解旋酶II产生的抗体特异性灭活。我们得出结论,DNA解旋酶II是uvrD基因产物。