Lysine-Phosphatidylcholine Adducts in Kringle V Impart Unique Immunological and Potential Pro-inflammatory Properties to Human Apolipoprotein(a)

作者：Celina Edelstein, Ditta Pfaffinger, Janet Hinman, Elizabeth R. Miller, Gregory M. Lipkind, Sotirios Tsimikas, Claes Bergmark, Godfrey S. Getz, Joseph Lee Witztum, Angelo M. Scanu · 发表于：Journal of Biological Chemistry · 年份：2003 · DOI：10.1074/jbc.m310425200 · 被引用次数：138 · 研究领域：Atherosclerosis and Cardiovascular Diseases、Cholesterol and Lipid Metabolism、Adipokines, Inflammation, and Metabolic Diseases

Lipoprotein(a), Lp(a), an athero-thrombotic risk factor, reacts with EO6, a natural monoclonal autoantibody that recognizes the phophorylcholine (PC) group of oxidized phosphatidylcholine (oxPtdPC) either as a lipid or linked by a Schiff base to lysine residues of peptides/proteins. Here we show that EO6 reacts with free apolipoprotein(a) apo(a), its C-terminal domain, F2 (but not the N-terminal F1), kringle V-containing fragments obtained by the enzymatic digestion of apo(a) and also kringle V-containing apo(a) recombinants. The evidence that kringle V is critical for EO6 reactivity is supported by the finding that apo(a) of rhesus monkeys lacking kringle V did not react with EO6. Based on the previously established EO6 specificity requirements, we hypothesized that all or some of the six lysines in human kringle V are involved in Schiff base linkage with oxPtdPC. To test this hypothesis, we made use of a recombinant lysine-containing apo(a) fragment, rIII, containing kringle V but not the protease domain. EO6 reacted with rIII before and after reduction to stabilize the Schiff base and also after extensive ethanol/ether extraction that yielded no lipids. On the other hand, delipidation of the saponified product yielded an average of two mol of phospholipids/mol of protein consistent with direct analysis of inorganic phosphorous on the non-saponified rIII. Moreover, only two of the six theoretical free lysine amino groups per mol of rIII were unavailable to chemical modifica...