Fractionation of non allelic histone variants by hydrophobic interaction chromatography

A method for the separation of non allelic histone variants is described. Sepharose-CL-4B was modified with dodecylamine by the carbonyldiimidazole-method. The matrix prepared contained 6 mumol/ml hydrophobic groups, C12-hydrocarbon chains. Proteins were loaded at high ionic strength and eluted from the column with an urea gradient. Histones H1, H2a and H2b from different mammalian tissues were loaded on the column and pure variants H1A, H1B, H2a1, H2a2, H2b1 and H2b2 were obtained by urea gradient elution. The recovery of the H2a-variants from calf thymus for example was 54% of H2a1 and 36% of H2a2.