Nucleic Acids Research, Vol 24, Issue 18 3527-3532, Copyright © 1996 by Oxford University Press
S Wold, E Crooke and K Skarstad
Fis protein participates in the normal control of chromosomal replication
in Escherichia coli. However, the mechanism by which it executes its effect
is largely unknown. We demonstrate an inhibitory influence of purified Fis
protein on replication from oriC in vitro. Fis inhibits DNA synthesis
equally well in replication systems either dependent upon or independent of
RNA polymerase, even when the latter is stimulated by the presence of HU or
IHF. The extent of inhibition by Fis is modulated by the concentrations of
DnaA protein and RNA polymerase; the more limiting the amounts of these,
the more severe the inhibition by Fis. Thus, the level of inhibition seems
to depend on the ease with which the open complex can be formed.
Fis-mediated inhibition of DNA replication does not depend on a functional
primary Fis binding site between DnaA boxes R2 and R3 in oriC, as mutations
that cause reduced binding of Fis to this site do not affect the degree of
inhibition. The data presented suggest that Fis prevents formation of an
initiation-proficient structure at oriC by forming an alternative,
initiation-preventive complex. This indicates a negative role for Fis in
the regulation of replication initiation.
ARTICLES
The Escherichia coli Fis protein prevents initiation of DNA replication from oriC in vitro
Department of Biophysics, Institute for Cancer Research, Montebello, Oslo, Norway.
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