Histone DNA complexes

Salt bridges between positively charged basic amino acid side chains of histones and the negatively charged DNA phosphates play a major role in stabilizing the DNA-histone complex. Indeed, treatment of chromatin with concentrated NaCl (1-2 m), which is known to disrupt electrostatic bonds, causes a complete dissociation of DNA and histone in the nucleohistone complex. 

In 1991, Luger et al. revealed by X-ray analysis the crystal structure of a natural DNA-histone complex. The X-ray structure shows in atomic detail how the histone protein octamer is assembled and how the base pairs of DNA are organized into a superhelix around it [74]. Since then this protein structure with cationic amino acids on the surface has acted as a model for the rational design of dendritic polymer-based gene vectors to mimic the globular shape of the natural histone complex [75-77]. 

J2. Jacob, L., Viard, J. P., Allenet, B., Anin, M. F., Slama, F. B., etal., A monoclonal anti-double-stranded DNA autoantibody binds to a 94-kDa cell-surface protein on various cell types via nucleosomes or a DNA-histone complex. Proc. Natl. Acad. Sci. USA 86, 4669—4673 (1989). 

Vibrational spectroscopy was used to study native chromatin as well as reconstituted DNA-histone complexes. IR (Liquier et al., 1979) and Raman spectra (Goodwin and Brahms, 1978 Savoie et al., 1985) show that the DNA in chromatin adopts a B type conformation. The important role adopted by the a helical parts of the histones in stabilizing the B conformation of histone-DNA complexes was demonstrated by IR (Taillandier et al., 1984b). Raman spectra of chromatin have made it possible to localize histone-DNA interactions in the minor groove and non-histone protein-DNA interactions in the major groove (Goodwin and Brahms, 1978). 

A conformational assessment of the DNA-histone complex and of its components at various pH values, ionic strengths and temperatures indicates that both components are mutually stabilized in the complex against denaturation (386). A recent CD study of histone-DNA complexes... 

Montminy, M. Transcriptional Activation Something New to Hang Your HAT On. Nature SST, 654-655 (1997). [Transcription in eukaryotes requires opening the DNA/histone complex, which can be controlled by acetylation.]... 

In eukaryotes, DNA supercoiling is an essential step in forming the DNA histone complexes of chromatin, and thus, in organizing the chromosome. DNA supercoiUng is controlled by a class of enzymes called topoisomerases [22]. We could demonstrate that the topoisomerase I molecule may be a target for ADP-ribosylation and that this covalent modification inactivates this enzyme involved in chromosome organization [23]. 

Table 1. Inhibition of poly(ADP-ribose) glycohydrolase by intercalators in the absence or presence of DNA-histone complexes ...

DNA-histone complexes (100 jug nil ) in the absence of intercalators, inhibit the enzyme activity by 39%... 

In the absence of DNA-histone complexes, ethacridine, proflavine, tilorone R10,556 DA, ellipticine and daunomycin inhibit the poly(ADP-ribose) glycohydrolase activity... 

Effect of DNA and DNA-Histone Complexes on the Inhibition of Poly(ADP-Ribose) Glycohydrolase by Intercalators. The inhibition of glycohydrolase activity by intercalators (proflavine, ellipticine, tilorone R10,556 DA) is relieved by the addition of 100 Mg mr calf thymus DNA (data not shown). This is probably due to the well-known binding of the intercalators to the added DNA. However, addition of DNA-histone complexes (100 jug nil" of each) slightly increases the inhibition of the glycohydrolase activity by those intercalators which are inhibitory (Table 1). These include ethacridine, tilorone R10,556 DA, eUipticine, daunomycin and proflavine. 

Fig. 1. Displacement of histone from DNA-histone complexes by intercalators 100 Mg of calf thymus DNA was added to 1 ml of 50 mM potassium phosphate buffer (pH 7.5) containing 100 Mg of histone and incubated for 15 min at 37°C. To remove the free histone, the DNA-histone complexes were washed (by centrifugation) three times with 50 mAf potassium phosphate buffer (pH 7.5), each time followed by 15 min incubation at 37°C. The washed complexes were suspended in 1 ml of 50 mM potassium buffer (pH 7.5) containing 200 mAf sodium chloride and 100 mA of the indicated intercalator. After incubation at 37° C for 30 min, the insoluble complexes were precipitated by centrifugation (8000 g 15 min, 4°C). 75 m1 of the supernatant was spotted on Whatman filter paper in 15 m1 aliquots. After drying, the filter was stained with 0.5% (w/v) Coo-massie blue in 7% (v/v) acetic acid, 30% (v/v) ethanol and destained with 7% acetic acid. 30% ethanol. 1 none, 2 proflavine, 3 ethacridine, 4 tilorone RIO,556 DA, 5 AMSA, 6 chloroquine, 7 ethidium bromide, 8-10 pure histone 0.5,1.5 and 2 Mg, respectively...

Those intercalators which are not inhibitory do not become inhibitors in the presence of DNA-histone complexes, except for ethidium bromide. Although in the absence of DNA-histone complexes, ethidium bromide is not inhibitory even at 200 fjM, in the presence of DNA-histone complexes, it inhibits the enzyme activity by 87% and 69% at 50 ijM and 20 [jM, respectively. 

The increased inhibition of the glycohydrolase in the presence of DNA-histone could be due to the formation of dye-DNA-histone complexes or the release of inhibitory histone from DNA by the intercalators. To test this suggestion the ability of these intercalators to release histone from DNA-histone complexes was studied. The method is described in the legend to Fig. 1 the displaced histone was separated from the insoluble DNA-histone complex by centrifugation. As shown in Fig. 1, there is a correlation between the ability of the intercalators to release histone from the complexes and the increase in their inhibition of the glycohydrolase activity in the presence of DNA-histone. AMSA (spot 5) and chloroquine (spot 6), which do not inhibit the enzyme activity in the presence of DNA-histone complexes, did not release histone from these complexes. However, proflavine (spot 2), ethacridine (spot 3) and tilorone (spot 4) which are inhibitors of the glycohydrolase and also display increased inhibition of the... 

The mechanism of inhibition by intercalators in the absence of DNA-histone complexes is not yet clear. However, the inhibition by ethacridine is probably due to its binding to the substrate, poly(ADP-ribose). This inference is drawn from the two observations that ethacridine inhibits the degradation of poly(ADP-ribose) not only by the glycohydrolase but also by the snake venom phosphodiesterase, and that ethacridine prevents the ethanol-acetate precipitation of poly(ADP-ribose). In the presence of DNA-histone complexes all inhibitory dyes produce a similar inhibition of the enzyme activity at equimolar concentrations. This suggests that the inhibition in these cases is due to the presence of the same amount of histones released by different intercalators. 

Isolated nuclei are highly permeable to histones, protamines and other biological macromolecules, whereas ATP and Na become tightly bound. The chief components of isolated and disrupted nuclei are DNA-histone complexes (nucleohistones), ribonucleic acids and poorly soluble acidic proteins (residual proteins). Nuclei also contain high concentrations of an arginase and an adenosine S -phosphata e of unknown function. 

Fig. 1. (left) Effect of poly(ADP-ribose) on DNA-histone complex formation. 40 ng samples of rat liver [ P]DNA were incubated with increasing concentrations of unlabeled poly(ADP-ribose) in 10 mM Tris-HCl (pH 7.0), 150 mM NaCl, 3 mM EDTA at22°C for 10 min. After addition of 20 ng histone HI, H3 or H4, respectively, and 10 min incubation in a final volume of 150 ul, the mixtures were placed on cellulose nitrate membrane filters, washed, and the amount of retained complexes measured. Other conditions as in (3). 

Clearly, the ionic bonding between DNA acidic-DNA phosphate group and the basic lysine residues of the histones constitutes a major mode of interaction between the two macromolecules, but ionic bonds are not the only type of bonds between DNA and histones. The finding that urea helps to dissociate the DNA-histone complex, especially in the case of the arginine-rich histones, suggests that hydrophobic bonding may be involved as well [74]. 

All types of histones are found in chromatin in equi-molecular amounts except for FI of which there is only half as much as of the other histones. We have seen that in the classical view, based on X-ray crystallography studies, the DNA histone complexes in chromatin are believed to form a supercoiled helical structure. Although it is certain that chromatin fibers are... 

It was demonstrated also that the acidic proteins stimulate synthesis in isolated chromatin fractions and increase the transcriptional activity of the DNA-histone complex (Frenster, 1965 Teng et al., 1970). These proteins promote the transcription of free DNA (Allfrey et al., 1972), and they determine transcriptional specificity. The accumulation of acidic nucleus proteins with a molecular weight of20,000-40,000 occurs during puff formation in Drosophila salivary gland chromosomes (Berendes, 1972). An important role in this process is played by phosphoproteins which demonstrate a high level of tissue specificity (Allfrey et al., 1972 Rickwood et al., 1972 Fig. 60). 

Category IV—Supramacromolecular (Exo-) those assemblies leading to precise, three-dimensional (3D) structure-controlled, noncovalently bound macromolecules. These supramacromolecular structures are derived from more complex, but precisely controlled macromolecular structures capable of information storage, expression, amplification, and use as functional/structural building blocks (e.g., protein folding, DNA-histone complexes, DNA expression, etc.). 

An attempt to represent the possible structure of the DNA-histone complex in repressed and active zones of DNA was undertaken by Frenster (1965) (Fig. 93). hi his model he postulates the existence of special RNA-derepressors containing anticodons for messenger RNA. 

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