Association of the Folded Chromosome with the Cell Envelope of Escherichia coli : Nature of the Membrane-Associated DNA

Abstract

Membrane-associated folded chromosomes isolated from Escherichia coli in the presence of spermidine sedimented at about 5,800S. The folded chromosome and the membrane fragment were each stable in the absence of the other; a 1,700S folded chromosome was obtained after removal of the membrane by a Sarkosyl treatment, and a 4,000S membrane fragment remained after digestion of the chromosomal DNA with deoxyribonuclease I. The interaction between the folded chromosome and the membrane fragment was stable, and, even when the DNA was unfolded, both components remained associated and cosedimented. The large frictional effect of the unfolded DNA reduced the sedimentation rate of the complex to about 2,000S. Partial removal of this unfolded DNA with restriction endonucleases caused the membrane fragments and the remaining associated DNA to sediment faster, at about 3,500S. The DNA remaining associated with the membrane fragments after restriction endonuclease treatment, about 4.5% of the total DNA when Eco RI was used, was indistinguishable from the DNA released from the membranes by three criteria: (i) DNA size distribution in agarose gels after electrophoresis, (ii) reassociation kinetics, and (iii) thermal elution from hydroxylapatite. This finding, that random DNA sequences rather than specific ones were responsible for the majority of the DNA-membrane interactions, argues against the folded chromosome's being a static structure with specific DNA sequences interacting with the cell envelope.