The chromatin‐associated protein H‐NS alters DNA topology in vitro

H‐NS is one of the two most abundant proteins in the bacterial nucleoid and influences the expression of a number of genes. We have studied the interaction of H‐NS with DNA; purified H‐NS was demonstrated to constrain negative DNA supercoils in vitro. This provides support for the hypothesis that H‐NS influences transcription via changes in DNA topology, and is evidence of a structural role for H‐NS in bacterial chromatin. The effects of H‐NS on topology were only observed at sub‐saturating concentrations of the protein. In addition, a preferred binding site on DNA was identified by DNase I footprinting at sub‐saturating H‐NS concentrations. This site corresponded to a curved sequence element which we previously showed, by in vivo studies, to be a site at which H‐NS influences transcription of the proU operon. When present in saturating concentrations, H‐NS did not constrain supercoils and bound to DNA in a sequence‐independent fashion, covering all DNA molecules from end to end, suggesting that H‐NS may form distinct complexes with DNA at different H‐NS:DNA ratios. The data presented here provide direct support for the hypothesis that H‐NS acts at specific sites to influence DNA topology and, hence, transcription.