Structural determinants in the C-terminal domain of apolipoprotein E mediating binding to the protein core of human aortic biglycan

Apolipoprotein (apo) E-containing high density lipoprotein particles were r eported to interact in vitro with the proteoglycan biglycan (Bg), but the d irect participation of apoE in this binding was not defined. To this end, w e examined the in vitro binding of apoE complexed with dimyristoylphosphati dylcholine (DMPC) to human aortic Bg before and after glycosaminoglycan (GA G) depletion. In a solid-phase assay, apoE DMPC bound to Bg and GAG-deplete d protein core in a similar manner, suggesting a protein-protein mode of in teraction. The binding was decreased in the presence of 1 M NaCl and was pa rtially inhibited by either positively (0.2 M lysine, arginine) or negative ly charged (0.2 nr aspartic, glutamic) amino acids. A recombinant apoE frag ment representing the C-terminal 10-kDa domain, complexed with DMPC, bound as efficiently as full-length apoE, whereas the N-terminal 22-kDa domain wa s inactive. Similar results were obtained with a gel mobility shift assay. Competition studies using a series of recombinant truncated apoEs showed th at the charged segment in the C-terminal domain between residues 223 and 23 0 was involved in the binding. Overall, our results demonstrate that the C- terminal domain contains elements critical for the binding of apoE to the B g protein core and that this binding is ionic in nature and independent of GAGs.