A human topoisomerase I cleavage complex is recognized by an additional human topisomerase I molecule in vitro.
Several recent studies have shown that human topoisomerase I (htopoI) can recognize various DNA lesions and thereby form a covalent topoisomerase I-DNA complex, which is known to be detrimental to cells. We have investigated whether htopoI recognizes another htopoI that is covalently trapped on a DNA substrate. For this purpose we created an artificial DNA substrate containing a specific topoisomerase I binding sequence, where the enzyme was trapped in the covalently bound form. We demonstrate that, in vitro, free htopoI stimulates the formation of an additional cleavage complex immediately upstream of the covalently bound topoisomerase I. The predominant distance between the two cleavage sites is 13 nt. In addition we find that these two enzymes may form direct protein-protein contacts and we propose that these may be mediated through the formation of a dimer by domain swapping involving the C-terminal and the core domains. Finally, we discuss the possibility that the double cleavage reaction may be the initial step for the removal of the recognized cleavage complex.
Søe
K
K
Institute of Molecular Biotechnology, Department of Biochemistry, Beutenbergstrasse 11, D-07745 Jena, Germany.
Dianov
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G
Nasheuer
H P
HP
Bohr
V A
VA
Grosse
F
F
Stevnsner
T
T
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D002166:Camptothecin / Q000494:pharmacology; D002384:Catalysis; D002460:Cell Line; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004264:DNA Topoisomerases, Type I / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D019286:Endopeptidase K / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D013379:Substrate Specificity; D020860:Subtilisin / Q000378:metabolism; D017186:Titrimetry; D059004:Topoisomerase I Inhibitors; D014357:Trypsin / Q000378:metabolism
D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D059004:Topoisomerase I Inhibitors; D004247:DNA; D014357:Trypsin; D020860:Subtilisin; D019286:Endopeptidase K; D004264:DNA Topoisomerases, Type I; D002166:Camptothecin