title: Mimicking lipid-binding-induced conformational changes in the human apolipoprotein E N-terminal receptor binding domain effects of low pH and propanol.
abstract: We studied the effects of n-propanol and pH on the structure of the apolipoprotein E3 N-terminal receptor binding domain, apo E3(1-191), to determine whether conditions similar to those occurring near lipid surfaces (decreased dielectric constant and pH) can mimic lipid-induced conformational changes in apo E3. The addition of 30% n-propanol, at pH 7, induces a conformational change in apo E3(1-191) as shown by changes in the intrinsic tryptophan fluorescence and by an increase in the Stokes radius of the majority of the protein from 3.0 to 4.1 nm, although the protein remains monomeric as shown by chemical cross-linking. These changes are accompanied by increased resistance to limited proteolysis with trypsin, chymotrypsin, subtilisin and endoproteinase glu-C, as is the case for apo E3(1-191) reconstituted into phospholipid/cholesterol lipid bicelles. Far and near UV circular dichroism showed that n-propanol increases the amount of calculated alpha-helical structure (42-65%) and alters the tertiary structure of the protein although not as much as when apo E3(1-191) is incorporated into lipid bicelles. In the absence of n-propanol, lowering the pH to 4.5 decreases the Stokes radius of the majority of the protein somewhat, with little effect upon the secondary and the tertiary structures. The addition of 30% n-propanol at pH 4.5 increases the Stokes radius of apo E3(1-191) from 2.2 to 5.0 nm, even more than at pH 7 (3.0-4.1 nm) although the protein still remains predominantly monomeric. There is increased resistance to limited proteolysis with endoproteinase glu-C. As assessed by far and near UV circular dichroism, the addition of 30% n-propanol at pH 4.5, in contrast to pH 7, markedly increases the alpha-helical structure and changes the tertiary structure of the protein similarly to that resulting from the incorporation of apo E3(1-191) into lipid bicelles. The results suggest that a combination of n-propanol and low pH in aqueous solutions may be useful as a simple model system for studying conformational changes in apo E3 similar to those, which occur upon interaction of the protein with lipids.
authors.affiliation: Centre de Génétique Moléculaire, Centre National de la Recherche Scientifique, Gif-sur-Yvette, Associé à l'Université Pierre et Marie Curie, Paris, France.
authors.forename: V
authors.initials: V
authors.lastname: Clément-Collin
authors.forename: A
authors.initials: A
authors.lastname: Leroy
authors.forename: C
authors.initials: C
authors.lastname: Monteilhet
authors.forename: L P
authors.initials: LP
authors.lastname: Aggerbeck
mesh_terms: D000433:1-Propanol / Q000494:pharmacology; D000817:Anilino Naphthalenesulfonates / Q000737:chemistry; D053318:Apolipoprotein E3; D001057:Apolipoproteins E / Q000737:chemistry*; D001665:Binding Sites; D002942:Circular Dichroism; D010450:Endopeptidases / Q000378:metabolism; D005453:Fluorescence; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D050356:Lipid Metabolism; D008081:Liposomes / Q000737:chemistry; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D010446:Peptide Fragments / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018110:Receptors, Lipoprotein / Q000378:metabolism; D013696:Temperature
chemical_list: D000817:Anilino Naphthalenesulfonates; D053318:Apolipoprotein E3; D001057:Apolipoproteins E; D008081:Liposomes; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D010446:Peptide Fragments; D018110:Receptors, Lipoprotein; C027132:1-anilino-8-naphthalenesulfonate; D000433:1-Propanol; D010450:Endopeptidases