Isolation of a non-histone chromosomal high mobility group protein from mouse liver nuclei by hydrophobic chromatography.

Abstract

A histone-binding polypeptide of 25,000 Mr (J2) in the saline-EDTA wash of mouse liver nuclei was purified to homogeneity in three steps: 1) ammonium sulfate fractionation, 2) hydrophobic chromatography using omega-amino butyl agarose, and 3) DEAE-cellulose chromatography. The isolation conditions were mild, and avoided extremes of pH and use of chaotropic reagents. The purified polypeptide was similar, although not identical, to calf thymus HMG-1 (high mobility group protein). The molecular weight, migration in two-dimensional polyacrylamide gels, and the amino acid sequence of the first 9 residues of the NH2-terminal region were identical for the two proteins. The amino acid analysis, however, indicated the mouse liver polypeptide was more acidic, with a lower content of lysine and higher contents of serine, glutamic acid, and aspartic acid. The mouse liver and calf thymus proteins differ in extraction from nuclei and solubility at neutral pH. The J2 polypeptide reacted with affinity-purified HMG-1 antibody by complement fixation, but always at a lower level than the homologous antigen purified with chaotropic reagents. We conclude the J2 polypeptide is an HMG-1-like protein. The intrinsic hydrophobicity of HMG-1 may be important in its functional interaction with DNA and histones.