Abstract
1. A procedure is presented for large scale fractionation of arginine-rich histones by exclusion chromatography. With this method, 2 to 3.5 g of histone Fractions 2a and 3 were fractionated. The two fractions contained the same histone components, but they differed in the amounts of each component. 2. In histone Fraction 2a, the glycine-rich, arginine-rich (GAR) histone accounted for approximately 70% of the protein present. This histone was obtained in a highly purified form after exclusion chromatography was repeated once, as shown by polyacrylamide gel electrophoresis, amino acid analysis, and peptide maps. Rechromatography of this product on Sephadex G-25 provided a GAR histone which was 96% pure. A quantity of GAR histone of 1 g was readily isolated by this procedure. 3. In histone Fraction 3, the arginine-rich, lysine-rich (AL) histone accounted for approximately 60% of the protein present. Rechromatography of the AL histone on Sephadex G-100 followed by rechromatography on Sephadex G-25 and Sephadex G-200 yielded a product approximately 96% pure. As in the case of the GAR histone, 1 g of the AL histone was readily obtained.
Highlights
Such studies to date have been difficult because of the lack of availability of a pure molecular species of histones. Recent studies in this laboratory have shown that arginine-rich histones of fetal thymes can be fractionated by exclusion chromatography [2]
One pass of histone Fraction 3 through a Sephadex G-100 column led to high yields of AL histone of similar purity to that obtained from the AR histones [2]
As with the exclusion chromatography of histone Fraction 2a, routinely 60 to 85% of the histone loaded was recovered from the column
Summary
1. A procedure is presented for large scale fractionation of arginine-rich histones by exclusion chromatography. 2. In histone Fraction 2a, the glycine-rich, arginine-rich (GAR) histone accounted for approximately 70% of the protein present. In histone Fraction 2a, the glycine-rich, arginine-rich (GAR) histone accounted for approximately 70% of the protein present This histone was obtained in a highly purified form after exclusion chromatography was repeated once, as shown by polyacrylamide gel electrophoresis, amino acid analysis, and peptide maps. Rechromatography of this product on Sephadex G-25 provided a GAR histone which was96% pure.
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