Association of apolipoprotein E with α2-macroglobulin in human plasma

Apolipoprotein (apo) E plays a central role in the transport of lipids among different organs and cell types, whereas alpha(2)-macroglobulin (alpha(2)M) is responsible for the binding and inactivation of plasma proteases, as well as the transport of various cytokines, growth factors, and hormones. In the present study, evidence is presented for direct binding of apoE with alpha(2)M in human plasma, based on the observation that two-dimensional non-denaturing gradient gel electrophoretic separation of plasma resulted in co-migration of apoE with alpha(2)M in a complex intermediate in size (18.5 nm in diameter) between low (LDL) and high density lipoproteins (HDL), ApoE associated with alpha(2)M could be immunoprecipitated from plasma with anti-human alpha(2)M antiserum. Purified apoE, labeled with I-125, bound to native and methylamine-activated alpha(2)M (alpha(2)M-MA) in vitro in a time- and concentration-dependent manner. ApoE bound to alpha(2)M-MA with greater affinity than alpha(2)M The binding of apoE to both alpha(2)M and alpha(2)M-MA did not depend on the presence of lipid. Ingestion of an oral fat load resulted in a reduction in the amount of apoE associated with alpha(2)M. Sphingomyelin vesicles and very low density lipoproteins (VLDL), but not phosphatidylcholine vesicles or HDL3, inhibited the in vitro binding of I-125-labeled apoE3 to alpha(2)M and alpha(2)M-MA. Binding of I-125-labeled apoE3 was also partially inhibited by an excess of platelet-derived growth factor and beta-amyloid protein, but not interferon-gamma, Subjects with an apoE 4/4 phenotype had less apoE associated with alpha(2)M in plasma than subjects with an apoE 3/3 or 2/2 phenotype, corresponding to reduced in vitro binding of apoE4 with alpha(2)M or alpha(2)M-MA. Although the functional significance of apoE binding to alpha(2)M remains to be determined, the present results demonstrate that: 1) apoE is non-covalently bound to alpha(2)M in human plasma, 2) alpha(2)M-MA has a greater capacity to bind apoE than alpha(2)M, 3) various proteins or lipoproteins known to bind apoE or alpha(2)M can potentially affect the interaction of apoE with alpha(2)M, and 4) association of apoE with alpha(2)M or alpha(2)M-MA is dependent on apoE phenotype.