title: Characterization of three human apolipoprotein E isoforms (E2, E3 and E4) expressed in Escherichia coli.
abstract: Apolipoprotein E is one of the apolipoproteins involved in cholesterol metabolism. Three major isoforms are present in men: E2, E3, E4 corresponding to the products of three alleles. They have different affinities for receptors and the epsilon 4 allele is a risk factor for cardiovascular diseases and more recently for Alzheimer's disease. We describe here the production, by heterologous expression in Escherichia coli, of the three apolipoprotein E isoforms for use in both research and clinical laboratories. By Surface Plasmon Resonance, the purified recombinant apolipoprotein E isoforms were able to recognize three monoclonal anti-human apolipoprotein E antibodies with affinity constants close to those of purified human apolipoprotein E. For receptor binding studies, the recombinant apolipoprotein E isoforms were associated with VLDL isolated from apolipoprotein E knockout mice. Although the association of the recombinant apolipoproteins E with the mouse VLDL was less efficient than that of human plasma apolipoprotein E3, the recombinant apolipoprotein E3 and apolipoprotein E4 complexes competed efficiently with 125I-labelled LDL for binding to the LDL receptor in J774 macrophages, whereas the recombinant apolipoprotein E2-VLDL complexes did not. These results suggest that the recombinant apolipoprotein E isoforms have biological properties similar to the human apolipoprotein E isoforms.
authors.affiliation: Centre du Médicament, Université de Nancy 1 H. Poincaré, URA CNRS 597, France.
authors.forename: A
authors.initials: A
authors.lastname: Barbier
authors.forename: A
authors.initials: A
authors.lastname: Visvikis
authors.forename: F
authors.initials: F
authors.lastname: Mathieu
authors.forename: L
authors.initials: L
authors.lastname: Diez
authors.forename: L M
authors.initials: LM
authors.lastname: Havekes
authors.forename: G
authors.initials: G
authors.lastname: Siest
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D053329:Apolipoprotein E2; D053318:Apolipoprotein E3; D053327:Apolipoprotein E4; D001057:Apolipoproteins E / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D015374:Biosensing Techniques; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D007536:Isomerism; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D011973:Receptors, LDL / Q000032:analysis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D053329:Apolipoprotein E2; D053318:Apolipoprotein E3; D053327:Apolipoprotein E4; D001057:Apolipoproteins E; D011973:Receptors, LDL; D011994:Recombinant Proteins