Lyamichev V, Brow M A, Dahlberg J E
Department of Biomolecular Chemistry, University of Wisconsin School of Medicine, Madison 53706.
Science. 1993 May 7;260(5109):778-83. doi: 10.1126/science.7683443.
Previously known 5' exonucleases of several eubacterial DNA polymerases have now been shown to be structure-specific endonucleases that cleave single-stranded DNA or RNA at the bifurcated end of a base-paired duplex. Cleavage was not coupled to synthesis, although primers accelerated the rate of cleavage considerably. The enzyme appeared to gain access to the cleavage site by moving from the free end of a 5' extension to the bifurcation of the duplex, where cleavage took place. Single-stranded 5' arms up to 200 nucleotides long were cleaved from such a duplex. Essentially any linear single-stranded nucleic acid can be targeted for specific cleavage by the 5' nuclease of DNA polymerase through hybridization with an oligonucleotide that converts the desired cleavage site into a substrate.
此前已知几种真细菌DNA聚合酶的5'核酸外切酶现在已被证明是结构特异性内切酶，可在碱基配对双链体的分叉末端切割单链DNA或RNA。切割与合成不偶联，尽管引物可显著加快切割速率。该酶似乎是通过从5'延伸的自由端移动到双链体的分叉处（切割发生的位置）来进入切割位点的。从这样的双链体上切割下了长达200个核苷酸的单链5'臂。本质上，任何线性单链核酸都可以通过与寡核苷酸杂交，将所需切割位点转化为底物，从而被DNA聚合酶的5'核酸酶靶向进行特异性切割。
