Shinder G, Gold M
Department of Medical Genetics, University of Toronto, Ontario, Canada.
J Virol. 1988 Feb;62(2):387-92. doi: 10.1128/JVI.62.2.387-392.1988.
The maturation and packaging of bacteriophage lambda DNA are under the control of the multifunctional viral terminase enzyme, which is composed of the protein products of Nu1 and A, the two most leftward genes of the phage chromosome. Terminase binds selectively to the cohesive end site (cos) of multimeric replicating lambda DNA and introduces staggered nicks to regenerate the 12-base single-stranded cohesive ends of the mature phage genome. The purified gpNu1 subunit of terminase forms specific complexes with cos lambda DNA. DNase I footprinting experiments showed that gpNu1 bound to three distinct regions near the extreme left end of the lambda chromosome. These regions coincided with two 16-base-pair sequences (CTGTCGTTTCCTTTCT) that were in inverted orientation, as well as a truncated version of this sequence. Bear et al. (J. Virol. 52:966-972,1984) isolated a mutant phage which contained a CG to TA transition at the 10th position of the rightmost 16-base-pair sequence, and this phage (termed lambda cos 154) exhibits a defect in DNA maturation when it replicates in Escherichia coli which is deficient in integration host factor. Footprinting experiments with cos 154 DNA showed that gpNu1 could not bind to the site which contained the mutation but could protect the other two sites. Since the DNA-packaging specificity of terminase resides in the gpNu1 subunit, these studies suggest that terminase uses these three sites as recognition sequences for specific binding to cos lambda.
噬菌体λ DNA的成熟和包装受多功能病毒末端酶的控制,该酶由噬菌体染色体最左侧的两个基因Nu1和A的蛋白质产物组成。末端酶选择性地结合多聚体复制型λ DNA的粘性末端位点(cos),并引入交错切口以再生成熟噬菌体基因组的12个碱基的单链粘性末端。纯化的末端酶gpNu1亚基与cos λ DNA形成特异性复合物。DNase I足迹实验表明,gpNu1与λ染色体最左端附近的三个不同区域结合。这些区域与两个反向的16碱基对序列(CTGTCGTTTCCTTTCT)以及该序列的一个截短版本一致。Bear等人(《病毒学杂志》52:966 - 972,1984)分离出一种突变噬菌体,其在最右侧16碱基对序列的第10位含有一个从CG到TA的转变,当这种噬菌体(称为λ cos 154)在缺乏整合宿主因子的大肠杆菌中复制时,其DNA成熟存在缺陷。用cos 154 DNA进行的足迹实验表明,gpNu1不能与含有突变的位点结合,但能保护其他两个位点。由于末端酶的DNA包装特异性存在于gpNu1亚基中,这些研究表明末端酶将这三个位点用作与cos λ特异性结合的识别序列。