The Resolution of Four Lysine-rich Histones Derived from Calf Thymus

Abstract

Abstract Histones rich in lysine were prepared by a procedure based on the direct extraction of calf thymus with aqueous trichloracetic acid. The protein obtained included what appeared to be degradation products, unless care was taken to isolate the histone rapidly (25 min) in the cold (0–3.5°) from thymus which had been frozen for transport to the laboratory. The protein preparations made according to this procedure appeared to be the same as other lysine-rich histone preparations described in the literature when examined by electrophoresis, chromatography, and amino acid analysis. These preparations seemed homogeneous in free boundary electrophoresis, and in zone electrophoresis in paper and starch gel. These preparations appeared homogeneous when examined by stepwise elution from carboxymethyl cellulose and when eluted from Amberlite IRC-50 by a gradient of barium acetate. Some resolution of these preparations was obtained by electrophoresis in polyacrylamide gel. This technique was especially powerful in detecting degradation products in the preparations. The most effective resolution of lysinerich histones, however, was obtained by chromatography on Amberlite IRC-50 with a gradient (7 to 14%) of guanidinium chloride in 0.1 m phosphate (pH 6.8). This method yielded three major fractions, one of which occasionally showed further partial resolution. Electrophoresis of the chromatographic fractions in polyacrylamide gel confirmed the presence of a total of four components in the lysine-rich histone prepared as noted above.