The QTK loop is essential for the communication between the N-terminal atpase domain and the central cleavage--ligation region in human topoisomerase IIalpha.
We have characterized a human topoisomerase IIalpha enzyme with a deletion of the conserved QTK loop, which extends from the transducer domain to the ATP-binding pocket in the GHKL domain. The loop has been suggested to play a role for interdomain communication in type II topoisomerases. The mutant enzyme performs only very low levels of strand passage, although it is able to cleave and ligate DNA as well as close the N-terminal clamp. Cleavage is nearly unaffected by ATP and ATP analogues relative to the wild-type enzyme. Although the enzyme is able to close the clamp, the clamp has altered characteristics, allowing trapping of DNA also in the absence of an ATP analogue. The enzyme furthermore retains intrinsic levels of ATPase activity, but the activity is not stimulated by DNA. Our observations demonstrate that the QTK loop is an important player for the interdomain communication in human topoisomerase IIalpha. First, the loop seems to play a role in keeping the N-terminal clamp in an open conformation when no nucleotide is present. Once the nucleotide binds, it facilitates clamp closure, although it is not essential for this event. The QTK loop, in contrast, is essential for the DNA-stimulated ATPase activity of human topoisomerase IIalpha.
Bendsen
Simon
S
Department of Molecular Biology, C. F. Moellers Alle, Building 1130, University of Aarhus, 8000 Arhus C, Denmark.
Oestergaard
Vibe H
VH
Skouboe
Camilla
C
Brinch
Marie
M
Knudsen
Birgitta R
BR
Andersen
Anni H
AH
D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000951:Antigens, Neoplasm / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism; D017931:DNA Primers; D004250:DNA Topoisomerases, Type II / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D008958:Models, Molecular; D016133:Polymerase Chain Reaction
D000951:Antigens, Neoplasm; D017931:DNA Primers; D004268:DNA-Binding Proteins; D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; D004250:DNA Topoisomerases, Type II