Hydrodynamic and physicochemical studies on bacteriophage PM2, intracellular and mature bacteriophage PM2 DNA, and a DNA-cleaving enzyme associated with the PM2 bacteriophage host

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1973

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Part I The sedimentation coefficient and intrinsic viscosity of nicked and closed circular PM2 bacteriophage DNA have been measured as a function of pH in the alkaline region. A gradual increase in the sedimentation coefficient, and a corresponding decrease in the intrinsic viscosity, are observed for the superhelical (closed) circle in the pH region from 10.5 to about 10.9. This has been tentatively interpreted in terms of the known dependence of sedimentation coefficient upon the number of superhelical turns. At slightly higher pH values, the curve passes through the minimum (sedimentation coefficient) and maximum (intrinsic viscosity) expected when the superhelical turns present at neutral pH are unwound by partial alkaline denaturation. Sedimentation studies of the relaxed (nicked) circular species have revealed the existence of DNA forms in the pH region from 11.27 to 11.37 which sediment considerably faster than the closed circle in the same region. These have been identified as partially denatured nicked circles in which varying fractions of the duplex structure have undergone alkaline denaturation, but strand separation has not yet occurred. Varying fractions of a slower species, either undenatured or completely denatured nicked circles, are also observed in some of these experiments. A corresponding result is observed in the intrinsic viscosity vs. pH curve. When nicked circular PM2 DMA is exposed to various alkaline pH's, rapidly neutralized, and sedimented at neutral pH, the expected sharp transition from native to denatured (strand-separated) molecules is seen. However, a very narrow pH range is noted in which native and denatured forms coexist in a single experiment. The above experiments carried out upon the closed form also reveal a narrow pH range in which the bulk of the transition from native closed circles to the collapsed cyclic coil takes place, in accord with an earlier study on a different DNA. This transi tion is shown never to be completely effected, however, as there is a fraction (7-8%) of the closed circles which renature to the native form, regardless of the alkaline pH employed. This same phenomenon was not observed in the case of artificially closed [lambda]b[subscript 2]b[subscript 5]c DNA circles. Possible explanations for some of the above results are discussed.[...]

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