Shlomai J, Linial M
J Biol Chem. 1986 Dec 5;261(34):16219-25.
Newly replicated duplex DNA minicircles of trypanosomal kinetoplast DNA are nicked in both their monomeric and catenated topological states, whereas mature ones are covalently sealed. The possibility that nicking may play a role during kinetoplast DNA replication by affecting the topological interconversions of monomeric DNA minicircles and catenane networks was studied here in vitro using Crithidia fasciculata DNA topoisomerase. An enzyme that catalyzes the nicking of duplex DNA circles has been purified to apparent homogeneity from C. fasciculata cell extracts. The native enzyme has a sedimentation coefficient of 6.8 S and was found to be a dimer with a protomer Mr = 60,000. Nicking of kinetoplast DNA networks by the purified enzyme inhibits their decatenation by the Crithidia DNA topoisomerase but has no effect on the catenation of monomeric DNA minicircles into networks. This differential effect on decatenation versus catenation is specific to the purified nicking enzyme. Random nicking of interlocked DNA minicircles has no detectable effect on the reversibility of the topological reaction. The potential role of Crithidia nicking enzyme in the replication of kinetoplast DNA networks in trypanosomatids is discussed.
锥虫动基体DNA新复制的双链DNA小环在其单体和连环拓扑状态下均有切口，而成熟的小环则是共价封闭的。本研究在体外使用纤细无鞭毛体DNA拓扑异构酶，探讨了切口是否可能通过影响单体DNA小环和连环网络的拓扑相互转换在动基体DNA复制过程中发挥作用。一种催化双链DNA环切口的酶已从纤细无鞭毛体细胞提取物中纯化至表观均一。天然酶的沉降系数为6.8 S，发现是一种二聚体，亚基Mr = 60,000。纯化的酶对动基体DNA网络的切口抑制了纤细无鞭毛体DNA拓扑异构酶对其解连环的作用，但对单体DNA小环连环形成网络没有影响。这种对解连环与连环的差异作用是纯化的切口酶所特有的。互锁DNA小环的随机切口对拓扑反应的可逆性没有可检测到的影响。本文讨论了纤细无鞭毛体切口酶在锥虫动基体DNA网络复制中的潜在作用。
