Topoisomerase II-mediated DNA cleavage: evidence for distinct regions of enzyme-DNA contacts.
To determine the specific interaction sites of topoisomerase II within the DNA region defined by the footprint of the enzyme, we have investigated the cleavage reaction on double-stranded DNA substrates containing nicks and deletions. Topoisomerase II-mediated cleavage of the DNA substrates is suicidal as the enzyme is unable to religate the cleaved DNA due to diffusion of the small nucleotides 5' to the cleavage position. Thus, suicidal cleavage is obtained with substrates having one, two or three nucleotides 5' to the cleavage position. The enzyme requires interaction with three distinct regions of double-stranded DNA for cleavage to occur, one region spanning the eight nucleotides located around the cleavage position and two distal regions each spanning approximately six nucleotides. A model is proposed, where these data are taken to imply that two distinct regions of interactions exist between each topoisomerase II subunit and its DNA substrate. The model is discussed in relation to the recently solved three-dimensional structure of yeast topoisomerase II.
Alsner
J
J
Department of Molecular and Structural Biology, University of Aarhus, Denmark.
Sorensen
H V
HV
Schmidt
V K
VK
Sorensen
B S
BS
Westergaard
O
O
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002417:Cattle; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004250:DNA Topoisomerases, Type II / Q000737:chemistry / Q000378:metabolism*; D008956:Models, Chemical; D008969:Molecular Sequence Data; D013379:Substrate Specificity
D004247:DNA; D004250:DNA Topoisomerases, Type II