Nucleoproteins. 2. Fractionation of deoxyribonucleoproteins into acidic proteins and lysine-rich histones associated with deoxyribonucleic acid

Abstract

An improved method of estimating amino acids, through their Cd2+ ion complexes, in nucleoproteins is described. DNA-protein complexes have been isolated from liver, spleen and tumour tissues by the action of 70 mM-benzoate or 15 mM-phenolphthalein diphosphate and phenol. The complexes from spleen and tumour tissues (isolated with 70mM-benzoate) could be fractionated by centrifuging. Most of the acidic protein sedimented with the DNA, whereas DNA associated with a lysine-rich histone remained in the supernatant solution. The proportion of lysine in the supernatant fraction after centrifuging complexes from liver was increased, but neither this increase nor the change in the alanine was sufficient to suggest the presence of a lysine-rich histone. A lysine-rich histone could be separated from the DNA-protein complex of rat-liver nuclei by centrifuging. Phenolphthalein diphosphate (15 mM) removed most of the residual protein from liver and spleen DNA but left a lysine-rich histone associated with the DNA but left a lysine-rich histone associated with the DNA from tumour tissues. These complexes could not be separated by centrifuging in the same manner as those prepared with 70 mM-benzoate. It is suggested that phenolphthalein diphosphate preferentially removes acidic proteins that form cross-links with DNA and render it more readily sedimentable.