The carboxyl terminus in apolipoprotein E2 and the seven amino acid repeat in apolipoprotein E-Leiden: role in receptor-binding activity.
Both apolipoprotein (apo) E2 and apoE-Leiden (tandem repeat of amino acids 121-127) are associated with type III hyperlipoproteinemia and bind defectively to low density lipoprotein receptors. Removing the carboxyl terminus of both variants (residues 192-299) increases receptor-binding activity, suggesting that the carboxyl terminus modulates activity. To identify the region(s) that modulated binding activity, we produced carboxyl-terminal truncations in apoE2 and apoE-Leiden (terminating at positions 191, 223, 244, and 272) and in apoE3 (terminating at positions 191, 223, and 244) and compared their receptor-binding activities as dimyristoylphosphatidylcholine (DMPC) discs. The results suggest that the entire carboxyl terminus up to residue 272, not a discrete smaller segment, is responsible for the modulation in apoE2. Intact apoE-Leiden and the 223 and 244 variants displayed similar activities (approximately 25% of apoE3's), but the 191 variant's activity was identical to that of intact apoE3. ApoE-Leiden and its truncated variants formed larger DMPC discs than did intact or truncated apoE3 or apoE2. These discs contained more apoE molecules than apoE3 discs, suggesting that the apparently normal binding activity of the apoE-Leiden 191 variant results from an increased number of apoE molecules and that the binding activity is actually defective. Direct comparison in a solid-phase assay revealed that the binding activity of the apoE-Leiden fragment was defective (51.4+/-9.4%). Thus, the defective binding of apoE-Leiden results from a direct effect of the seven amino acid repeat on receptor-binding activity rather than from an indirect effect operating through the carboxyl terminus as previously believed.
Dong
L M
LM
Gladstone Institute of Cardiovascular Disease, University of California, San Francisco, 94141-9100, USA.
Innerarity
T L
TL
Arnold
K S
KS
Newhouse
Y M
YM
Weisgraber
K H
KH
D053329:Apolipoprotein E2; D053318:Apolipoprotein E3; D001057:Apolipoproteins E / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D004134:Dimyristoylphosphatidylcholine; D014644:Genetic Variation; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D017433:Protein Structure, Secondary; D011973:Receptors, LDL / Q000097:blood* / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism
D053329:Apolipoprotein E2; D053318:Apolipoprotein E3; D001057:Apolipoproteins E; D011973:Receptors, LDL; D011993:Recombinant Fusion Proteins; C052720:apolipoprotein E3 (Leidein); D004134:Dimyristoylphosphatidylcholine