Studies on Deoxyribonucleoprotein Structure

Abstract

1. The addition of tRNA to chromatin gel (initial insoluble deoxyribonucleoprotein preparation) in the presence of 1 mM MgC1, or of 40 mM NaCl results in the transfer of the very lysinerich histone (Fl) and of some nonhistone proteins to the tRNA, whereas all other histones remain in the deoxyribonucleoprotein. This result provides a new general method for the mild removal of histone F1 from the chromatin gel. The addition of tRNA to chromatin gel in the absence of Mga+ and Na+ results not only in the rapid removal of histone F1 and of some nonhistone proteins from the chromatin gel, but also in the slow transfer of histones F2a2 and F2b to the tRNA. A similar pattern of histone redistribution is observed when the tRNA (or DNA) is added to the soluble deoxyribonucleoprotein preparation obtained by mechanical disruption of the chromatin gel.2. There is no protein redistribution in the above-mentioned soluble deoxyribonucleoprotein preparation (in the absence of exogeneous free polynucleotides).3. The addition of free DNA or tRNA to the soluble deoxyribonucleoprotein preparation, obtained by treatment of the chromatin gel with 2 M urea, allows one to remove quantitatively histones F1, F2a2 and F2b from the deoxyribonucleoprotein, leaving the initial distribution of the arginine-rich histones (F2al and F3) on the endogeneous DNA unchanged.4. The redistribution of proteins (and in particular of histones) occurs in the soluble deoxyribonucleoprotein preparation obtained by urea treatment even in the absence of any exogeneous polynucleotides suggesting that conservation of the initial distribution of proteins in this soluble deoxyribonucleoprotein preparation is not complete. 16.close to the initial native deoxyribonucleoprotein, i.e. that no redistribution of proteins occurs during or after the isolation procedure.Jensen and Chalkley [12] have shown that the introduction of free DNA into the soluble deoxyribonucleoprotein preparation (obtained by mechanical disruption ofthe chromatin gel) results in the transfer ofa part of the lysine-rich histone to exogeneous DNA. However, no further analysis of this phenomenon was carried out. I n the present paper the question is studied in more detail in order to characterize the features of the interactions between deoxyribonucleoprotein and free DNA or RNA and also between Merent deoxyribonucleoprotein molecules in the soluble preparations.The following questions were analysed. (a) What is the pattern of protein redistribution in the soluble deoxyribonucleoprotein preparations containing exogeneous free DNA or RNA? (b) Does the possibility of the transfer of proteins from the chromatin gel (initial insoluble deoxyribonucleoprotein preparation) to the free DNA or RNA exist ? (c) Does any redistribution of proteins in the soluble deoxyribonucleoprotein preparations occur?

Changes in Chromatin Properties after Partial Extraction of Non‐histone Proteins

“…1 , The densitometer scans were retraced and the areas under the curves were cut and weighed to obtain relative amounts of the histone fraction. The resul; were expressed as mean percentage k standard error from 10 independent isolations of whole chromatin and 5 independent isolations of tRNA- 131 histone has been shown by several authors [2, 3,8]. Consistent with these reports, the solubility of saltextracted chromatin (i.e., HI-depleted chromatin extracted in 0.6 M NaCI) (Fig.…”

“…Transfer RNA has been employed previously by Ilyn et al [8] for the selective removal of histone fractions from chromatin. It has been noted, however, that treatment of chromatin with tRNA leads not only to extraction of various histones but also to the removal of considerable portion of non-histone proteins [S].…”

“…The procedure for the extraction of proteins from chromatin with tRNA was based on the study of Ilyin et al [8]. 15-fold excess of tRNA (with respect to DNA concentration of the chromatin) in 5 mM Tris-HCI pH 8.0, 1 mM MgC12, 0.1 mM PhMeS03F was added with stirring to the viscous solution of chromatin in 1 mM Tris-HC1 pH 8.0, 0.1 mM PhMeS03F (1.5 mg/ml chromatin DNA).…”

Chromatin architecture: investigation of a subunit of chromatin by dark field electron microscopy.

“…2a). Two distinct types of interparticle fibers were discovered, those of [20][21][22][23][24][25][26][27][28][29][30] A width, presumably containing a single duplex DNA molecule, and those of A width, often formed by the coalescence of two of the thinner fibers.…”

Studies of Interaction between Histone F2b and DNA from Normal and Exposed to X-radiation Calf Lymph Nodes

“…Cell nuclei were obtained by a modified sucrose method (Majkowska, Lemierz and Klyszejko-Stefanowicz 1971) . The purity of this histone fraction was checked by electrophoresis in polyacrylamide gels (Ilyin, Varshavsky, Mickelsaar and Georgev 1971), modified for comparative purposes .…”