No abstract

The conformational changes of histones and DNA during the thermal denaturation of native and partially histone-depleted nucleoprotein (DNA . protein) was studied by following the variations of absorbance and circular dichroism with temperature.All the measurements were made in 1 mM NaCl, 0.2 mM EDTA pH 5.6. Absorbance was measured a t 260 nm and circular dichroism (C.D.) a t 280 nm and 227 nm, in order to follow the conformational changes of DNA and histones separately. Besides the prcmelting effect (which is not observed with native DNA * protein), the second maximum which takes place, in the 280-nm C.D. melting profile, a t a temperature close to 65 "C, can be interpreted as a change from the C form to the B form of DNA.If non-histone proteins are assumed to have no effect upon the structure of DNA, the height of the second maximum varies almost linearly with the histone content except, may be, in the case of Fzal. Any histone would thus be equivalent to induce conformational changes of DNA. This change is always preceded and probably triggered by the cooperative disruption of a large amount of &-helical regions of the histones which in their new tertiary structure are no more able to maintain the DNA in a C conformation.In the model of the structural changes of DNA . protein with temperature, which is tentatively proposed, histone-histone interactions would play a role as important as histone-DNA interactions.The deoxyribonucleoprotein extracted from the nuclei of eukaryotic cells is a complex of DNA with basic proteins (histones) of limited heterogeneity and various acidic proteins. Two possible functions have been suggested for these proteins. It can be said roughly that acidic proteins are involved in the control mechanism of genetic regulation, but histones are necessary to stabilize the condensed structure of DNA in the chromatin.There is now some evidence suggesting that the native nucleoprotein exists in the form of a supercoil. The evidence is based on studies of fibres or gels using X-ray diffraction [ given wavelength A . relative to a 1-em optical path; C.D., circular dichroism; d Ot 1, difference between ellipticities measured a t t "C and 25 "C, respectively, and for a given wavelength I ; protein/DNA, ratio (w/w) of protein to DNA in native or partially stripped nucleoprotein. Definition. A,,, unit, the quantity of material contained in 1 ml of a solution which has an absorbance of 1 a t 260 nm, when measured in a 1-cm pathlength cell. hydrodynamic properties of nucleohistones and the wide-angle X-ray scattering curves also give evidence of a compact conformation in dilute solution [5-9]. According to circular dichroism studies, the secondary structure of DNA in chromatin has been found to be different from that of isolated DNA in solution [8,10--161. It is unknown so far whether the changes in the secondary structure of DNA are correlated with the changes in the tertiary structure. The precise manner in which the histone fractions interact with DNA in the chromatin is still unknown.I n order t...

Spectroscopic studies (nuclear magnetic resonance, circular dichroism and infrared) have been carried out on chicken erythrocyte histone H 5 and on three peptides cleaved therefrom: 1-31, 32-197 and 58-197. It is shown that at ionic strengths above 0.1M part of the H 5 molecule takes up a globular conformation containing 14% a helix but no p sheet structure. Several details of the circular dichroism and nuclear magnetic resonance spectra indicate that the globular region is located in the N-terminal half of the molecule and this proposal is supported by the observation that the peptide 32-197 is largely incapable of folding and the peptide 59-197 is completely incapable of folding. Structural similarities and differences between histone H 5 and histone H 1 are discussed.The nucleated erythrocytes of birds, reptiles, amphibians and fish contain a very basic histone, H5, that has not been found in other tissues [l -51. H 5 largely, but not completely, replaces H 1 and the two histones are not dissimilar in amino acid composition and molecular weight [4,6 -81. A significant difference between the two histones is that H 5 contains 11 % arginine whilst H 1 contains only 2%. In this respect H 5 resembles the 4 1 histones of marine invertebrate sperm (in particular sea urchins) which totally replace the H I and also contain about 11% arginine in addition to roughly 25% lysine 191. Early observations of the occurrence and composition of H 5 resulted in the suggestion that its function is the total suppression of the genome in the fully mature erythrocyte [lo]. Detailed studies on anemic chickens have subsequently shown, however, that H 5 can be detected even in erythroblasts (when a wide range of protein synthesis is taking place), and does not appear suddenly at the mature erythrocyte stage when protein synthesis is finally terminated [I 1 -151. The function of H 5 in relation to genome suppression is, therefore, not as simple as originally thought. H 5 nevertheless is closely related to H I [I61 and a number of experiments have implicated H 1 in the condensation of diffuse chromatin [17 -191. In particular the state of chromatin is thought to depend on the degree of phosphorylation of H 1 [20,21] and recently it has been shown that H 5 is phosphorylated and dephosphorylated in viuo [22]. There is at present no evidence to Abbreviations. NMR, nuclear magnetic resonance; CD, circular dichroism.suggest why H 1 should be replaced in large part by H 5 and the present structural investigation of H 5 was carried out to point out the similarities and differences between H 5 and H 3 . Related studies on H 1 have already been published [23,24]. Hydrodynamic studies have shown that H 5 does not aggregate at high ionic strength [8,28] and thereby resembles H 1 and not the remaining four histone fractions. The monomeric state of H 1 has led to proposals that its function is quite separate from that of the other histones and this may also be true for H5. In chicken erythrocytes the stoichiometry of histones H 5 and H 1 together, wi...