Preferential relaxation of supercoiled DNA containing a hexadecameric recognition sequence for topoisomerase I

In prokaryotes, the degree of supercoiling of DNA can profoundly influence the use of specific promoters (for review, see ref. 1). In eukaryotes, a variety of indirect observations suggest that DNA topology has a similar importance in proper gene expression2–4. Much attention has therefore been focused on the cellular proteins that control DNA supercoiling, among which are the enzymes topoisomerase I and II (for review see refs 5 and 6). A hexadecameric sequence functions as a strong attraction site for topoisomerase I7. Here we report that the interaction of topoisomerase I with this sequence motif is highly specific, because a single base-pair substitution prevents strand cleavage and thereby catalytic activity at the sequence. Thus, supercoiled DNA containing the recognition sequence is relaxed preferentially by topoisomerase I compared to a control, but no difference in the relaxation rate is observed for supercoiled DNA carrying the mutated sequence. The preference for the recognition sequence seems to be an intrinsic property of all eukaryotic type I topoisomerases8, suggesting that the interaction might be important in a fundamental biological process.