Project Details


Our long-range goal is to understand how chromosome structure affects DNA
replication and gene expression. The present proposal focuses on how DNA
topoisomerases and supercoiling affect gene expression. We have shown
that two topoisomerases are involved in determining the level of DNA
supercoiling in bacteria and we now propose to address three questions:
1) can special features of DNA secondary structure be identified near
genes whose expression is affected by superhelical density? 2) does DNA
gyrase bind to DNA more frequently near actively transcribing genes, and
if so, where are the binding sites? and 3) is the expression of a large
number of genes affected by changes in supercoiling? Our general strategy
involves perturbing the normal level of supercoiling inside cells using
topoisomerase mutations, cloned topoisomerase genes and inhibitors of
gyrase. DNA secondary structures will be detected by their sensitivity to
cleavage by single strand-specific nucleases, gyrase locations will be
identified by oxolinic acid-induced DNA cleavage and expression of
specific genes by radioactive labeling of proteins that are then
identified by gel electrophoresis. Understanding the function of DNA
supercoiling is one of the central issues in bacterial chromosome biology
and we expect the information gained from this study to form part of the
knowledge base necessary to understand and hopefully cure, genetic
Effective start/end date7/1/853/31/86


  • National Institutes of Health


  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)

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