An unusual type IB topoisomerase from African trypanosomes.
African trypanosomes are ancient eukaryotes that cause lethal disease in humans and cattle. Available drugs are inadequate and the need for new therapeutic targets is great. Trypanosoma brucei and related pathogens differ strikingly from higher eukaryotes in many aspects of nucleic acid structure and metabolism. We find yet another example of this in their unusual DNA topoisomerase IB. Type IB topoisomerases relieve the supercoils that accumulate during DNA and RNA synthesis, and are of considerable importance as the target for antitumor camptothecins. Dozens of type IB topoisomerases sequenced from eukaryotes, bacteria, and pox viruses are all encoded by a single gene that predictably contains a highly conserved DNA binding domain and C-terminal catalytic domain, linked by a nonconserved hydrophilic region. We find that topoisomerase IB in T. brucei is encoded by two genes: one for the DNA-binding domain and a second for the C-terminal catalytic domain. In keeping with this, highly purified fractions of native T. brucei topoisomerase IB catalytic activity contain two proteins, of 90 and 36 kDa. The native enzyme is conventional in its Mg2+-independence, ability to relax positive and negative supercoils, and inhibition by camptothecin. Camptothecin promotes the formation of a covalent complex between 32P-labeled substrate DNA and the small subunit. This unusual structural organization may provide a missing link in the evolution of type IB enzymes, which are thought to have arisen over time from the fusion of two independent domains. It also provides another basis for the design of selectively toxic drug candidates.
Bodley
Annette L
AL
Division of Clinical Pharmacology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Chakraborty
Asit K
AK
Xie
Suji
S
Burri
Christian
C
Shapiro
Theresa A
TA
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D002166:Camptothecin / Q000494:pharmacology; D020134:Catalytic Domain; D002845:Chromatography; D004247:DNA / Q000378:metabolism; D004264:DNA Topoisomerases, Type I / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics*
D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D004247:DNA; C475926:DNA topoisomerase IB, Trypanosoma brucei; D004264:DNA Topoisomerases, Type I; D002166:Camptothecin