A new method for isolating the nonidentical protein subunits of human plasma alpha-lipoprotein.

Human plasma alpha lipoprotein (alphaLP) was totally delipidated by gel filtration on Sephadex LH-20 in a medium of 2 butanol:acetic acid:H(2)O, 4:1:5. The resulting alpha protein (alphaP) exhibited two major bands, labeled C and D, on acrylamide-gel electrophoresis in 5.0 M urea at pH 8.8 or 4.0. Minor bands labeled A and B, also present, were shown to be aggregates of C which form when the latter is lyophilized. The C and D components were isolated in pure form from alphaP (prepared by LH-20 chromatography of alphaLP) by gel filtration of this protein on Sephadex G-200 in a medium of 1.0 N acetic acid: the C component emerged with a distribution coefficient (K(d)) of 0.4, and the D component with a coefficient of 0.7. From each 100 mg of alphaP, 68 mg of C and 22 mg of D were isolated. 3 mg of a minor fraction with K(d) 0.1, containing A and B components as well as C, were also obtained. D but not C reacts with rabbit antiserum to human alphaLP. C and D differ substantially in content of arginine, histidine, (1/2)-cystine, isoleucine, and tryptophan.