The structure and properties of histone F2a comprising the heterologous group F2a1and F2a2studied by13C nuclear magnetic resonance

Clark, V. M.; Lilley, David M.J.; Howarth, Oliver W.; Richards, B. M.; Pardon, J. F.

doi:10.1093/nar/1.7.865

Cited by 20 publications

(21 citation statements)

References 16 publications

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“…This is in keeping with our previous finding (1) that both pairs are required together with DNA to generate the x-ray diffraction pattern characteristic of chromatin. It is also in keeping with the work of D'Anna and Isenberg (20) and Clark et al (21) but not with the suggestion of Hyde and Walker (22) that the pairs F2A1,FS and F2A2,F2B do not interact.…”

Section: Methodssupporting

confidence: 86%

An octamer of histones in chromatin and free in solution.

Thomas¹,

Kornberg²

1975

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

651

207

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Crosslinking with dimethyl suberimidate reveals a chain of histone octamers in chromatin. The octamer can be isolated free in solution at high ionic strength and pH. The identification of dimers formed by crosslinking reveals two or more contacts of each histone with others within the octamer. The molecular weight (110,000) and pattern of dissociation of the octamer are compatible with the composition (F2A1)2(F3)2(F2A2)g(F2B)2. The four main types of histone associate in pairs in solution: histones F2A1 and F3 constitute a tetramer (1-3) of composition (F2A1)2(F3)2, and histones F2A2 and F2B a mixture of oligomers, possibly dimers or short polymers (1,(4)(5)(6). The finding of a histone tetramer led to a model for chromatin structure (7) (8) containing 0.2 mM NaEDTA, pH 7. F1 was removed by the tRNA exchange method (13) in 40 mM NaCl/3 mM NaEDTA, pH 8.Procedure for Crosslinking. Buffers were 70 mM sodium phosphate for pH 8, ionic strength 0.2; 60 mM NaCl/100 mM sodium borate for pH 9, ionic strength 0.1; and 1.95 M NaCI/100 mM sodium borate for pH 9, ionic strength 2.0. Chromatin in buffer was treated at 23' with Me2Sub, freshly dissolved at 20 mg/ml in buffer, or with dithiobis(succinimidyl propionate), freshly dissolved at 50 mg/ml in dimethylformamide. Solutions were dialyzed against 0.2 mM phenylmethylsulfonyl fluoride/0. 1 mM NaEDTA, pH 7 and freeze-dried for analysis in gels.Sodium Dodecyl Sulfate (NaDodSO4)/Polyacrylamide Gel Electrophoresis. NaDodSO4/5% polyacrylamide tube gels (14), 0.4 cm X 7 cm, were run at 6 mA per gel. NaDodSO4/polyacrylamide slab gels contained 18% acrylamide and were run according to Laemmli (15) with three modifications: the concentration of Tris buffer in the separating gel was increased to 0.75 M; the ratio acrylamide: N,N'-methylene bis-acrylamide was lowered (16) to 30:0.15; and the electrode buffer contained 0.05 M Tris, 0.38 M glycine, and 0.1% NaDodSO4. The slabs were 0.15 cm thick and either 15 cm or 30 cm long. The short gels were run at 30 mA for about 6 hr and the long ones at 4 W or less for about 24 hr, until the bromophenol blue tracking dye had reached the bottom. Tubes and slabs were fixed for at least 1 hr in methanol/acetic acid/water (5:1:5), stained in 0.1% (w/v) Coomassie brilliant blue in the same solvent, and destained by diffusion at 370 in 5% methanol/7.5% acetic acid. Migration was from top to bottom or left to right in all gels and densitometer traces shown.RESULTS AND DISCUSSION Native chromatin Chromatin prepared by a method involving limited nuclease digestion is native, whereas chromatin prepared by conventional methods involving shear is not (12). In the experiments reported here we have either worked directly with the product of nuclease digestion (a mixture of chromatin fragments containing DNA of weight-average size 1600 base pairs, which we refer to as "1600 base-pair chromatin") or with pure chromatin fragments (monomer, dimer, trimer, etc. of the repeating unit) obtained from the mixture by fractionation on a sucrose gradient.Most of ...

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Section: Methodssupporting

confidence: 86%

An octamer of histones in chromatin and free in solution.

Thomas¹,

Kornberg²

1975

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

651

207

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“…Mobile C a carbons are consistent only with random coil peptides; ' (iii) Relative mobility should be deduced from relaxation times. Previous n.m.r, investigations [4,16,23,24] of histones have relied upon differences in linewidth to estimate mobilities. This cannot be used as a rigorous test of backbone mobility since observed resonances from histones are generally composed of a number of partially overlapping resonances and hence v½ does not reflect 7"2 alone.…”

Section: Resultsmentioning

confidence: 99%

“…Histones interact with each other and with DNA [1][2][3][4][5][6] ;both kinds of association are essential for the integrity of chromatin. Current models for the structure of chromatin are based upon a periodic repeating unit in which a segment of DNA comprising about 140-160 base pairs is folded with a packing ratio of 7 or less into a 'bead' containing eight histone molecules, two each of F2A1, F3, F2A2 and F2B [7,8].…”

Section: Introductionmentioning

confidence: 99%

“…Using ~SC nuclear magnetic resonance (n.m.r.) we have studied the conformation of individual histones when alone in solution [4,16] and in the grouping found in chromatin. In this paper we show that the histones in the group containing two molecules each of histones F2A1 and F3 have random coil N-terminal peptides which are ideally equipped to bind to DNA.…”

Section: Introductionmentioning

confidence: 99%

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The existence of random coil N‐terminal peptides — ‘tails’ — in native histone complexes

Lilley¹,

Howarth²,

Clark³

1976

FEBS Letters

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“…It has been possible both to determine which regions of the histones are involved in homologous interactions and also to show that the aggregation process does not affect the secondary structure of the histones. The original I3C studies of histones F2Al and F2A2 (Clark et al, 1974) have been extended and similar studies made with histones F2B and F3.…”

mentioning

confidence: 98%

Investigation of the conformational and self-aggregational processes of histones using hydrogen and carbon-13 nuclear magnetic resonance

Lilley¹,

Howarth²,

Clark³

et al. 1975

Biochemistry

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Histone self-aggregation processes have been studied by 13C and 1H nuclear magnetic resonance (NMR) as a function of ionic strength and protein concentration. Thus has led to a model involving apolar aggregation between structured regions of these molecules. This analysis supports the validity of the acquistion of conformational data on histones by the simulation of 13C NMR spectra at high concentration. Solution conformations for histones F2B and F3 are presented.

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The structure and properties of histone F2a comprising the heterologous group F2a₁and F2a₂studied by¹³C nuclear magnetic resonance

Cited by 20 publications

References 16 publications

An octamer of histones in chromatin and free in solution.

An octamer of histones in chromatin and free in solution.

The existence of random coil N‐terminal peptides — ‘tails’ — in native histone complexes

Investigation of the conformational and self-aggregational processes of histones using hydrogen and carbon-13 nuclear magnetic resonance

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