Apoprotein (E--A-II) complex of human plasma lipoproteins. I. Characterization of this mixed disulfide and its identification in a high density lipoprotein subfraction.

AM, = 46,000 apoprotein designated as the apo(E-AII) complex has been isolated from the d < 1.006 fraction of human type III hyperlipoproteinemic plasma and from the d = 1.063 to 1.125 fraction of normal human plasma. The apo(E-A-II) complex was stable to treatment with 4 M guanidine, 8 M urea, and 2% sodium dodecyl sulfate but was dissociated into two subunits following reduction with /Smercaptoethanol or dithiothreitol. A M, = 37,000 subunit was shown to be identical with the E apoprotein by co-electrophoresis on polyacrylamide gels, immunochemical identity, amino acid analysis, and isoelectric focusing. The second subunit with a molecular weight of 8,500 was shown by coelectrophoresis, immunoreactivity, and amino acid analysis to be identical with the A-II apoprotein. In addition to being present in the d < 1.006 fraction of normal and type III hyperlipoproteinemic subjects, the apo(E-A-II) complex occurred as a major apoprotein constituent in a minor subclass of the high density lipoproteins (HDL) of normal human plasma. This subclass, referred to as HDL-I, was isolated by Geon-Pevikon preparative block electrophoresis from the d = 1.063 to 1.125 fraction and was found to be distinctively different from the main HDL constituent (referred to as HDL-II) of this ultracentrifugal fraction. All subclasses of the HDL contained the A-I and A-II apoproteins; however, the presence of the (E-A-II) and E apoproteins in the HDL-I distinguished this subclass from the HDL-II and HDLI (d = 1.125 to 1.21). Treatment of HDG I with a reducing agent (&mercaptoethanol) converted the apo(E-A-II) complex to the E and A-II apoproteins in the native particle without altering other chemical properties of these lipoproteins. As described in the accompanying paper (Innerarity, T. L., Mahley, R. W., Weisgraber, K. H., and Bersot, T. P. (1978) J. Biol. Chem. 253, 6289-6295), most, if not all, of the high affinity binding of HDL (d = 1.063 to 1.21) to low density lipoprotein receptors on the surface of human fibroblasts was accounted for by the activity of the HDL-I. Furthermore, the conversion of the apo(E-A-II) complex to the E apoprotein in uitm markedly enhanced the binding activity of the HDL-I. The possible interconversion between the apo(E-A-II) complex and the E and A-II apoproteins in uiuo and its importance in lipoprotein metabolism remains to be determined.