The domain organization and properties of individual domains of DNA topoisomerase V, a type 1B topoisomerase with DNA repair activities

被引:24
作者
Belova, GI
Prasad, R
Nazimov, IV
Wilson, SH
Slesarev, AI
机构
[1] Fidel Syst Inc, Gaithersburg, MD 20879 USA
[2] Russian Acad Sci, MM Shemyakin & Yu A Ovchinnikov Inst Bioorgan Che, Moscow 117871, Russia
[3] NIEHS, Struct Biol Lab, NIH, Res Triangle Pk, NC 27709 USA
关键词
D O I
10.1074/jbc.M110131200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Topoisomerase V (Topo V) is a type IB (eukaryotic-like) DNA topoisomerase. It was discovered in the hyperthermophilic prokaryote Methanopyrus kandleri and is the only topoisomerase with associated apurinic/apyrimidinic (AP) site-processing activities. The structure of Topo V in the free and DNA-bound states was probed by limited proteolysis at 37 degreesC and 80 degreesC. The Topo V protein is comprised of (i) a 44-kDa NH2-terminal core subdomain, which contains the active site tyrosine residue for topoisomerase activity, (ii) an immediately adjacent 16-kDa subdomain that contains degenerate helix-hairpin-helix (HhH) motifs, (iii) a protease-sensitive 18-kDa HhH "hinge" region, and (iv) a 34-kDa COOH-terminal HhH domain. Three truncated Topo V polypeptides comprising the NH2-terminal 44-kDa and 16-kDa domains (Topo61), the 44-, 16-, and 18-kDa domains (Topo78), and the COOH-terminal 34-kDa domain (Topo34) were cloned, purified, and characterized. Both Topo61 and Topo78 are active topoisomerases, but in contrast to Topo V these enzymes are inhibited by high salt concentrations. Topo34 has strong DNA-binding ability but shows no topoisomerase activity. Finally, we demonstrate that Topo78 and Topo34 possess AP lyase activities that are important in base excision DNA repair. Thus, Topo V has at least two active sites capable of processing AP DNA. The significance of multiple HhH motifs for the Topo V processivity is discussed.
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页码:4959 / 4965
页数:7
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