title: The lipid-associated conformation of the low density lipoprotein receptor binding domain of human apolipoprotein E.
abstract: Apolipoprotein E (apoE) is a 34-kDa exchangeable apolipoprotein that regulates metabolism of plasma lipoproteins by functioning as a ligand for members of the LDL receptor family. The receptor-binding region localizes to the vicinity of residues 130-150 within its independently folded 22-kDa N-terminal domain. In the absence of lipid, this domain exists as a receptor-inactive, globular four-helix bundle. Receptor recognition properties of this domain are manifest upon lipid association, which is accompanied by a conformational change in the protein. Fluorescence resonance energy transfer has been used to monitor helix repositioning, which accompanies lipid association of the apoE N-terminal domain. Site-directed mutagenesis was used to replace naturally occurring Trp residues with phenylalanine, creating a Trp-null apoE3 N-terminal domain (residues 1-183). Subsequently, tyrosine residues in helix 2, helix 3, or helix 4 were converted to Trp, generating single Trp mutant proteins. The lone cysteine at position 112 was covalently modified with N-iodoacetyl-N'-(5-sulfo-1-naphthyl)ethylenediamine, which serves as an energy acceptor from excited tryptophan residues. Fluorescence resonance energy transfer analysis of apoE N-terminal domain variants in phospholipid disc complexes suggests that the helix bundle opens to adopt a partially extended conformation. A model is presented that depicts a tandem arrangement of the receptor-binding region of the protein in the disc complex, corresponding to its low density lipoprotein receptor-active conformation.
authors.affiliation: Department of Biochemistry, University of Alberta, Edmonton, Alberta T6G 2S2, Canada.
authors.forename: C A
authors.initials: CA
authors.lastname: Fisher
authors.forename: V
authors.initials: V
authors.lastname: Narayanaswami
authors.forename: R O
authors.initials: RO
authors.lastname: Ryan
mesh_terms: D001057:Apolipoproteins E / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D006801:Humans; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011973:Receptors, LDL / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D001057:Apolipoproteins E; D011973:Receptors, LDL