Human apolipoprotein E4 domain interaction. Arginine 61 and glutamic acid 255 interact to direct the preference for very low density lipoproteins.
Human apolipoprotein (apo) E contains an amino- and a carboxyl-terminal domain, which are connected by a hinge region (approximately residues 165 to 215). The interaction of the two domains has been suggested to be responsible for the apoE4-binding preference for very low density lipoproteins (VLDL). In the absence of this interaction in apoE3, the preference is for high density lipoproteins (HDL). To exclude the possibility that the interaction of apoE with other apolipoproteins on the native particles may contribute to the isoform-specific preferences, VLDL-like emulsion particles were incubated with apoE, and the lipid-bound apoE was separated from free apoE on a Superose 6 column. The apoE4 bound more effectively to these particles than did apoE3, indicating that the apoE4 preference for VLDL is due not to interactions with other apolipoproteins but to an intrinsic property of apoE4, likely related to domain interaction. Previously, arginine 61 was shown to be critical for the isoform preferences, suggesting that it interacted with an acidic residue(s) in the carboxyl terminus. Substitution of arginine 61 with lysine did not alter the preference of apoE4 for VLDL, demonstrating that a positive charge rather than a specific requirement for arginine is critical for domain interaction. To identify the acidic residue(s) in the carboxyl terminus interacting with arginine 61, the six acidic residues (244, 245, 255, 266, 270, and 271) in a region known to be important for both lipoprotein association and isoform-specific preferences were substituted individually with alanine in apoE4. Only substitution of glutamic acid 255 altered the preference of apoE4 from VLDL to HDL, indicating that this was the sole residue in the carboxyl terminus that interacts with arginine 61. The participation of the hinge region in domain interaction was examined with internal deletion mutants. Deletion of the residues 186-202 or 186-223, representing major portions of the hinge region, had no effect on the apoE4 preference for VLDL. This suggests that the hinge region may act as a spacer that connects the two domains. Further deletion into the carboxyl-terminal domain (to residue 244) results in a loss of apoE4 VLDL binding. These studies establish that interaction of arginine 61 and glutamic acid 255 mediates apoE4 domain interaction.
Dong
L M
LM
Gladstone Institute of Cardiovascular Disease, University of California, San Francisco, California 94141-9100, USA.
Weisgraber
K H
KH
D053327:Apolipoprotein E4; D001057:Apolipoproteins E / Q000235:genetics / Q000378:metabolism*; D001120:Arginine / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism*; D006801:Humans; D008079:Lipoproteins, VLDL / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017384:Sequence Deletion
D053327:Apolipoprotein E4; D001057:Apolipoproteins E; D008079:Lipoproteins, VLDL; D018698:Glutamic Acid; D001120:Arginine