A comprehensive fractionation procedure for avian erythrocyte histones.

Abstract

Abstract : A fractionation procedure for avian erythrocyte histones was devised to give the six known, reproducible histone fractions from a single nuclear preparation, with limited manipulation and little wastage. The proposed method includes selective, two-step acid extraction, followed by cation-exchange chromatography and exclusion chromatography. The fractions were identified by amino acid analysis and by starch gel electrophoresis. The method is flexible in scale and permits a continuous balance of compositions and activities in metabolic experiments. Factors contributing to the relative extractability of histones from chromatin in acid were considered. The essential role of this step was the separation of erythrocyte-specific, serine-rich histone V, and arginine-rich histones III, IV, which otherwise would be eluted together from Amberlite CG-50 by guanidinium chloride. Two other arginine-rich components, more easily eluted in chromatograms of other tissues, were absent from the usual extracts of mature erythrocytes. However, similar components were produced from histones III, IV by treatment with beta-mercaptoethanol. The conditions for good resolution of sub-fractions of moderately lysine-rich histones and of arginine-rich histones by exclusion chromatography on Bio-Gel P-60 and P-30 were examined. Reversible adsorption appeared to be more significant than pore size of the gel; the sulphates of arginine-rich histones were resolved better than the chlorides. (Author)