The molecular structure of nucleohistone (DNH)

Richards, B. M.; Pardon, J. F.

doi:10.1016/0014-4827(79)90519-6

Cited by 141 publications

(41 citation statements)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…I n other words if any of these fractions are specifically involved, then they can only carry out their supercoiling function in the presence of f2a2. The necessary requirement for f2a2 would be consistent with X-ray studies on reconstituted nucleoprotein [ 2 ] . On the other hand none of the fractions need play it unique role in supercoiling and it may be sufficient simply for a certain proportion of the histone, regardless of composition, to be dissociated for the supercoiling to be lost.…”

Section: Discussionsupporting

confidence: 53%

“…On the other hand none of the fractions need play it unique role in supercoiling and it may be sufficient simply for a certain proportion of the histone, regardless of composition, to be dissociated for the supercoiling to be lost. Diffraction patterns of fibres from which the protein has been removed by dialysis against 1.0 M NaCl show that the reflections attributed to supercoiling are still present [ 2 ] , whereas in solution we have shown that the viscosity has increased, the…”

Section: Discussionmentioning

confidence: 68%

See 1 more Smart Citation

The Structure of Nucleohistone

Henson

Walker

1971

European Journal of Biochemistry

View full text Add to dashboard Cite

The viscosity of histone-depleted nucleohistone has been measured a t high and low ionic strength. Native nucleohistone a t low ionic strength and f l -depleted nucleohistone a t both low and high ionic strength all have the same intrinsic viscosity of 10.0 dl/g. This implies that these molecules are not flexible but behave in solution as rigid particles. On the assumption that they are rods they have a weight-average length of 1160 f 120 nm and diameter 5.6 & 0.3 nm.The DNA is compressed in the rod to give an increase in mass per unit length for the DNA of 5.0 0.5.By contrast the intrinsic viscosity of nucleohistone depleted of f l , f2a2 and 75O/, of F2b and F3, varies with ionic strength in the manner typical of a flexible polyelectrolyte. The shapes of the sedimentation boundaries measured at high ionic strength of depleted nucleohistone show that the dissociation of histone from DNA takes place so that all molecules are depleted to the same extent. It thus has a conformation intermediate between that of native nucleohistone and DNA.Polyacrylamide gel electrophoresis of histones removed by 1 .O M NaCl using a technique capable of resolving all five main histone fractions, shows that if one fraction only is responsible for supercoiling nucleohistone it must be f2a2.There is some evidence that in fibres of nucleohistone the DNA is held in the form of a compact supercoil by the histones [l]. The hydrodynamic properties of nucleohistone suggest that the compact conformation is also present in solutions of the complex as both the viscosity and the frictional coefficient of nucleohistone increased sharply as histone was dissociated in salt [3]. These changes were interpreted as an increase in asymmetry of the molecule resulting from the loss of the supercoiled structure. However, the hydrodynamic studies were carried out in solutions of low ionic strength and it can be argued that the increased electrostatic charge on the complex consequent on the removal of histones could give rise to polyelectrolyte effects which might account €or all the changes observed. I n this study we present viscosity data measured a t high ionic strength where polyelectrolyte effects are negligible. New data on the selectivity of the salt dissociation of histones, together with the viscosity observations a t high ionic strength have enabled us to make a more complete interpretation of the properties of the nucleohistone molecule in solution. EXPERIMENTAL PROCEDUREThe preparation and characterisation of deoxyribonucleohistone and deoxyribonucleohistone partially depleted of histone have been described else-1 Eur. 3. Biochem.. Vol.22 where [3]. Histone-depleted samples are referred to by the concentration of NaCl used to dissociate the protein. Thus 0.6 deoxyribonucleohistone has been depleted in 0.6 M NaCl. Dissociated histone was separated from the nucleohistone complex using gel exclusion chromatography. Relative viscosities and sedimentation coefficients were measured as described previously [3]. The procedure for polyacrylamide gel...

show abstract

Section: Discussionsupporting

confidence: 53%

Section: Discussionmentioning

confidence: 68%

The Structure of Nucleohistone

Henson

Walker

1971

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…1, model II) and contain 200 base pairs of DNA, giving a packing ratio of about 7:1. The length of the unit in this model comes from x-ray diffraction, which shows a 100 A repeat in the fiber direction (5,6), and the width comes from electron micrographs showing fibers about 100 A thick (7,8). The amount of DNA per unit is derived from biochemical results (9,10) and has been substantiated by studies involving brief micrococcal nuclease digestion (11).…”

mentioning

confidence: 99%

Electron microscopy of defined lengths of chromatin.

Finch¹,

Noll²,

Kornberg³

1975

Proc. Natl. Acad. Sci. U.S.A.

139

View full text Add to dashboard Cite

Defined lengths of chromatin were prepared by brief digestion with micrococcal nuclease and fractionation in a sucrose gradient. A length containing a given number of 200 base pair repeating units appeared as the same number of 100 A beads in the electron microscope. The distance between beads within a length was small, usually less than about 20 A. Several independent lines of work have led to the idea of beadlike units in chromatin, but there is disagreement over the spacing and DNA content of the beads. Two models have been put forward. One stems from a particulate appearance of chromatin fibers in the electron microscope, observed by Olins and Olins (1) The aim of the work described here was to determine the relationship between the bead observed in the electron microscope and the 200 base pair repeating unit inferred from biochemical evidence. Counts of beads in chromatin containing defined lengths of DNA, in particular SV40 chromatin (12) and complexes formed in vitro between adenovirus DNA and histones (13), have suggested that the bead and the biochemical unit are the same. Chromatin containing defined lengths of DNA can also be obtained by brief digestion with micrococcal nuclease and fractionation in a sucrose gradient (11). Here we report the counts of beads in such fractions. In the course of this work we have also made measurements of inter-bead distances in an effort to decide between the extended and compact models described above. MATERIALS AND METHODSChromatin (A26o = 46) was prepared as described (ref. 15, with the use of 110 units/ml of micrococcal nuclease for 2 min at 370 and lysis in half the stated volume of 0.2 mM NaEDTA) and 0.8 ml was layered on each of five 36.3 ml isokinetic sucrose gradients, containing 1 mM NaEDTA, pH 7, with ct = 5%, c, = 28.8% and Vm = 33 ml (16). The gradients were centrifuged for 22 hr at 27,000 rpm and 40 in a Beckman SW27 rotor, and analyzed with the use of a turbulence-free flow cell (Molecular Instruments Co, Evanston, Ill.). Corresponding fractions of the gradients (Fig. 2) were pooled, and dialyzed overnight against 1 mM NaEDTA, pH 7. The purity at this stage, determined as described in the text, was 95, 87, 78, and 67% for monomer, dimer, trimer and tetramer fractions. An aliquot (3 ml) of each fraction was further purified on a second sucrose gradient identical to the first.For electron microscopy, the purified fractions were dialyzed against 0.5 mM NaEDTA, pH 7, adjusted to the required ionic strength by the addition of ammonium acetate, and fixed if required in 5% formaldehyde, pH 7, for 5-10 min at 300. Drops of solutions were applied to thin carboncoated grids and the excess solution was withdrawn with an edge of filter paper.

show abstract

“…6, stage 8) Pairing of homologous chromosomes proceeds in three steps (11): (i) Approximate alignment to within 300 nm. (ii) Precise point-to-point pairing, at a distance of 100 nm, by the synaptinemal complex.…”

mentioning

confidence: 99%