Ethanol precipitation analysis of thymus histone

Abstract

An analytical ethanol precipitation technique, similar to 's salting-out procedure, was used for the characterisation of whole thymus histone and the products obtained by preparative ethanol fractionation. The analysis was carried out at —5° C and pH 6.5. Whole histone prepared according to et al., and to et al. (the product prepared according to the latter having been dialysed against phosphate, pH 6.5) appeared not to differ essentially; it contained among other substances a component precipitating at about 7% ethanol. If dialysis or storage at pH values approaching the neutral point were avoided in the et al. method, this component was no longer observed. By means of ultracentrifugation it was proved to be an aggregate, the formation of which cannot be prevented in the et al. method.

The et al. method was simplified and the HCl concentration determined at which the histone is completely split off from the nucleohistone. Ethanol precipitation analysis revealed the presence of two components in the whole histone prepared in this way, if the above-mentioned measures are taken to prevent aggregate formation. Component I, representing 80% of the whole histone was precipitated between 13 and 17% ethanol; component II, representing 20% of the whole histone, never completely precipitated upon increasing the ethanol concentration. The latter component was dissociated from the nucleohistone at much lower acidity than the former.

Component I (S20 = 1.7 ± 0.1) appeared to be arginine-rich, Component II (S20 = 0.9 ± 0.1) lysine-rich. Ethanol fractionation is recommended for preparing the arginine-rich and lysine-rich histone fractions.