Open AccessContacts between DNA gyrase and its binding site on DNA: features of symmetry and asymmetry revealed by protection from nucleases.
Author(s) -
Alan Morrison,
Nicholas R. Cozzarelli
Publication year - 1981
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.78.3.1416
Subject(s) - dna gyrase , dna supercoil , circular bacterial chromosome , dna , replisome , exonuclease iii , base pair , biology , exonuclease , topoisomerase , dna clamp , binding site , microbiology and biotechnology , biochemistry , dna replication , dna polymerase , escherichia coli , gene , polymerase chain reaction , reverse transcriptase
DNA gyrase supercoils DNA by passing one DNA segment through another by means of a reversible double-strand break at specific DNA sites. We determined the nucleotide sequence of two highly preferred gyrase binding sites and analyzed the grip of gyrase on the DNA by using protection from nuclease attack. The DNA-breakage site of gyrase was centered in about 50 base pairs (bp) of DNA that was completely protected from DNase I and flanked by DNA regions cut at average intervals of 9.9 bases. The same pattern of protection from DNase I was observed with topoisomerase II', an enzyme that shares structural homology with gyrase. The gyrase site of DNA breakage was off-center in the 140 bp of DNA protected from exonuclease III digestion. ATP or inhibitors of gyrase had little specific effect on DNase I protection. On addition of a nonhydrolyzable analogue of ATP, previously stable barriers to exonuclease III were invaded and new barriers appeared. We discuss a detailed model uniting these results with previous data on gyrase structure and mechanism.