The partition locus of plasmid pSC101 is a specific binding site for DNA gyrase.
A protein in extracts of Escherichia coli that specifically binds the stabilizing par sequence of pSC101 was identified as DNA gyrase. The purified enzyme protects par against digestion by DNase I and exonuclease III. Competition assays demonstrate that gyrase has a 40-fold higher affinity for the 100-bp par sequence than for nonspecific DNA and that par is the major gyrase-binding site in pSC101 derivatives used in this and other studies. Within par, AT-rich sequences occur with a pronounced 10-bp periodicity that is shifted by 5 bp from a similar periodicity of GC-rich sequences. As judged by DNase I digestion, the GC sequences are exposed on the outside of the DNA wrapped around gyrase. The data suggest that the site-specificity of DNA gyrase may be partly determined by the bendability of the DNA. A 4-bp deletion that interferes with Par function in vivo also reduces the affinity for gyrase in vitro. However, a deletion of par causes little reduction in superhelical density in vivo. We conclude that DNA gyrase, while involved in the Par function, may not affect plasmid stability through its supercoiling activity or by an influence on DNA replication.
Wahle
E
E
Department of Biochemistry, Stanford University School of Medicine, CA 94305.
Kornberg
A
A
D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004278:DNA, Superhelical / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010957:Plasmids; D012045:Regulatory Sequences, Nucleic Acid; D012093:Replicon
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004278:DNA, Superhelical; D004250:DNA Topoisomerases, Type II