Zhang H L, Malpure S, DiGate R J
Molecular and Cell Biology Program, University of Maryland at Baltimore 21201, USA.
J Biol Chem. 1995 Oct 6;270(40):23700-5. doi: 10.1074/jbc.270.40.23700.
The binding of DNA topoisomerase III (Topo III) to a single-stranded DNA substrate containing a strong cleavage site has been examined. The minimal substrate requirement for Topo III-catalyzed cleavage has been determined to consist of 7 bases; 6 bases 5' to the cleavage site and only 1 base 3' to the site. Nuclease P1 protection experiments indicate that the enzyme also binds to its substrate asymmetrically, protecting approximately 12 bases 5' to the cleavage site and only 2 bases 3' to the cleavage site. A catalytically inactive mutant of Topo III shows the same protection pattern as the active polypeptide, indicating that Topo III is a site-specific binding protein as well as a topoisomerase. Consistent with this view, an oligonucleotide containing a cleavage site is a more effective inhibitor and is bound more efficiently by Topo III than an oligonucleotide without a cleavage site.
已对DNA拓扑异构酶III（Topo III）与包含强切割位点的单链DNA底物的结合进行了研究。已确定Topo III催化切割的最小底物需求由7个碱基组成；切割位点5'端有6个碱基，3'端仅有1个碱基。核酸酶P1保护实验表明，该酶也以不对称方式结合其底物，在切割位点5'端保护约12个碱基，在切割位点3'端仅保护2个碱基。Topo III的催化失活突变体显示出与活性多肽相同的保护模式，表明Topo III不仅是一种拓扑异构酶，也是一种位点特异性结合蛋白。与这一观点一致的是，含有切割位点的寡核苷酸是一种更有效的抑制剂，并且与没有切割位点的寡核苷酸相比，Topo III对其结合效率更高。
