摘要：SummaryGene silencing in bacteria is mediated by chromatin proteins, of whichEscherichia coliH-NS is a paradigmatic example. H-NS forms nucleoprotein filaments with either one or two DNA duplexes. However, the structures, arrangements of DNA-binding domains (DBDs), and positions of DBD–DNA contacts in linear and bridged filaments are uncertain. To characterize the H-NS DBD contacts that silence transcription by RNA polymerase, we combined ·OH footprinting, molecular dynamics, statistical modeling, and DBD mapping using a chemical nuclease (Fe2+-EDTA) tethered to the DBDs (TEN-map). We find that H-NS DBDs contact DNA at indistinguishable locations in bridged or linear filaments and that the DBDs vary in orientation and position with ∼10-bp average spacing. Our results support a hemi-sequestration model of linear-to-bridged H-NS switching. Linear filaments able to inhibit only transcription initiation switch to bridged filaments able to inhibit both initiation and elongation using the same irregularly spaced DNA contacts.Graphical abstractDisplay OmittedHighlights•Tethered-nuclease mapping (TEN-map) of H-NS DNA-binding domains detects DNA contacts•Bridged and linear H-NS filaments use the same DNA contact sites•H-NS-DNA contacts are unevenly spaced with ∼10-bp average separation•AT-steps, minor groove width, and electrostatic potential best predict contact sitesBiological sciences; Molecular biology; Molecular mechanism of gene regulation
