Bacterial DNA topoisomerase I can relax positively supercoiled DNA containing a single-stranded loop.
Using heteroduplex molecules formed from a pair of plasmids, one of which contains a small deletion relative to the other, it is shown that bacterial topoisomerase I can relax a positively supercoiled DNA if a short single-stranded loop is placed in the DNA. This result supports the postulate that the specificity of bacterial DNA topoisomerase I for negatively supercoiled DNA in its relaxation reaction derives from the requirement of a short single-stranded DNA segment in the active enzyme-substrate complex. Nucleolytic and chemical probing of complexes between bacterial DNA topoisomerase I and heteroduplex DNA molecules containing single-stranded loops ranging from 13 to 27 nucleotides in length suggests that the enzyme binds specifically to the region containing a single-stranded loop; the site of DNA cleavage by the topoisomerase appears to lie within the single-stranded loop, with the enzyme interacting with nucleotides on both sides of the point of cleavage.
D001345:Autoradiography; D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D004264:DNA Topoisomerases, Type I / Q000378:metabolism*; D004270:DNA, Circular; D004278:DNA, Superhelical / Q000378:metabolism*; D004587:Electrophoresis, Agar Gel; D004926:Escherichia coli / Q000201:enzymology; D008837:Micrococcus / Q000201:enzymology; D010957:Plasmids; D013379:Substrate Specificity
D001426:Bacterial Proteins; D004270:DNA, Circular; D004278:DNA, Superhelical; D004264:DNA Topoisomerases, Type I