Immunochemistry of Human Very Low Density Lipoproteins: Apolipoprotein C-III

Apolipoprotein C-III (apoC-III) is a major protein constituent of human plasma very low density lipoproteins (VLDL) and a minor constituent of high density lipoproteins (HDL). The apoprotein is a single polypeptide chain of 79 amino acids and occurs in several forms differing only in their content of sialic acid. In the present report a quantitative radioimmunoassay (RIA) has been developed in order to study the immunochemical properties of apoC-III. Two individual rabbit antisera were used. It was observed that the COOH-terminal half (residues 41-79) of apoC-III containing 1 mol of sialic acid (apoC-III₁) contains the antigenic reactive region(s), whereas the NH₂-terminal half of the molecule was unable to react with anti-apoC-III antibodies. When the positively charged lysines of apoC-III₁ were chemically modified by acetic anhydride, the immunoreactivity was decreased by ~40% as measured by RIA in one rabbit antiserum. However, there was no decrease of immunoreactivity in another rabbit antiserum, suggesting that with the former antibody several populations exist directed at different regions of the apoprotein molecule. In addition, the immunoreactivity of apoC-III₁-phospholipid complexes was investigated and was found to be indistinguishable from that of apoC-III₁. Since the conformation of the apoprotein has drastically changed upon the addition of dimyristoylphosphatidylcholine (DMPC), this finding indicates that the gross conformational change of apoC-III₁ does not affect the immunochemical properties and that the antigenic reactive sites are probably located at the surface in apoC-III₁-DMPC complexes. The immunoreactivity of apoC-III was also found to be approximately the same in HDL or VLDL as that of the delipidated apoHDL or apoVLDL. Thus, the antigenic sites of apoC-III must be fully exposed on the surface of the lipoproteins.