Deoxyribonucleic acid (DNA) of Escherichia coli was found to be attached to the cell membrane at about 20 points. This was determined by fractionation of X-irradiated cells with the M band (magnesium-Sarkosyl crystals) technique. The number of attachment points was computed from the relationship between the amount of DNA in M bands and the number of double-strand breaks introduced by the X-ray treatment. The number of attachment points was decreased fourfold by treatment of cells with rifampin. This effect was apparently due to the action of the drug on ribonucleic acid (RNA) polymerase since the drug did not affect a mutant whose RNA polymerase is resistant to rifampin. This suggests that there may be two classes of attachment points of DNA on the membrane, some of which are removed by rifampin treatment and some which are not. Rifampin treatment also resulted in the uncondensing of isolated nucleoids and in an axial appearance of the nucleoids in ultrathin sections. The results suggest that RNA polymerase plays a role, direct or indirect, in maintaining the structure of the bacterial nucleoid and in some of its attachment to the membrane.

Lysozyme was immobilized by two different methods in two different ways in order to obtain a preparation with an activity as high as possible toward a macromolecular substrate. The enzyme was bound as a Schiff base to a silicate carrier by using oxidized dextrans of different lengths as spacer and also was bound to controlled pore aminoglass via pyridino-4-aldehyde and BrCN. The latter preparations had activities up to 2.5% of the free lysozyme.

No abstract

Until recently it has been assumed that the factors being responsible for the condensation of the DNA in the nucleoid of Escherichia coli are destroyed by rifampicin because it has been impossible to obtain folded chromosomes from cells treated with this inhibitor. I n this paper i t is shown by viscosity and sedimentation measurements that unfolding of the DNA does not take place during the process of the cell lysis as it should be predicted from this assumption, but is occurring distinctly afterwards. Since the observed unfolding process is too slow to be caused simply by molecular movements it is concluded that i t is brought about by the action of salts or detergents of the lysis medium. The structure of the nucleoid is still intact in viva despite inhibition of RNA synthesis by rifampicin.The chroniosonial DNA of Escherichia coli is a single circular molecule of the length of 1 p i (CAIRNS 1963) which is highly folded up in the cell. This folding is due to internal factors of the chromosonie and not to external forces like pressure exerted by the contents packed into the bacterial cell. Therefore, the DNA can be isolated in a folded or condensed state, the so-called nuclear body or nucleoid. But if the cells had been treated with rifampicin, an inhibitor of initiation of RNA synthesis, no nuclear bodies could be prepared, and the DNA was obtained in an extended gelatinous form (DWORSKY and SCHAECHTER 1973, PETTIJOHN et al. 1973). It was concluded from these experiments that inhibition of transcription destroys the factors being responsible for DNA condensation.However, this conclusion is not quite correct because the lysis method used (which will be called method A, and is described in "Materials and methods") niay be LOO aggressive for rifampicin treated cells. So it was found that (1) after treatment of E. coli with rifampicin the lysate occasionally was non-viscous at first, and the DNA sediniented with a velocity comparable to that of nuclear bodies from untreated cells (DWORSKY 1974) that is, the DNA was not unfolded in all preparations, and ( 2 ) that it is possible by using other lysis procedures to obtain chromosomes in a folded state also from E. coli which has been incubated with rifampicin (DWORSKY 1975).One of these procedures which seemed to be optimal (it will be called method B)uses lower concentrations of sodium chloride and a mixture of three detergents, among them cetyltriniethylammonium bromide. However, it cannot be excluded that a t least one of both lysis media A or B causes a considerable alteration of the nuclear bodies because particles from cells not treated with an antibiotic differ much in respect to their sensitivity towards ribonuclease and more concentrated salt solutions depending on the method of preparation. Proteins playing an essential role in DNA condensation may be dissociated by higher concentrations of detergents or salts as they are used in method A. Otherwise, structures of the nucleoid may be rearranged in such a way by niethod B that unfolding is inhibited during l...