A two-module region of the low-density lipoprotein receptor sufficient for formation of complexes with apolipoprotein E ligands.
The low-density lipoprotein (LDL) receptor transports two different classes of cholesterol-carrying lipoprotein particles into cells: LDL particles, which contain a single copy of apolipoprotein B-100 (apoB-100), and beta-migrating very low-density lipoprotein (beta-VLDL) particles, which contain multiple copies of apolipoprotein E (apoE). The ligand-binding domain of the receptor lies at its amino-terminal end within seven adjacent LDL-A repeats (LA1-LA7). Although prior work clearly establishes that LA5 is required for high-affinity binding of particles containing apolipoprotein E (apoE), the number of ligand-binding repeats sufficient to bind apoE ligands has not yet been determined. Similarly, uncertainty exists as to whether a single lipid-activated apoE receptor-binding site within a particle is capable of binding to the LDLR with high affinity or whether more than one is required. Here, we establish that the LA4-5 two-repeat pair is sufficient to bind apoE-containing ligands, on the basis of binding studies performed with a series of LDLR-derived "minireceptors" containing up to four repeats. Using single chain multimers of the apoE receptor-binding domain (N-apoE), we also show that more than one receptor-binding site in its lipid-activated conformation is required to bind to the LDLR with high affinity. Thus, in addition to inducing a conformational change in the structure of N-apoE, lipid association enhances the affinity of apoE for the LDLR in part by creating a multivalent ligand.
Fisher
Carl
C
Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, 77 Avenue Louis Pasteur, Boston, Massachusetts 02115, USA.
Abdul-Aziz
Dunia
D
Blacklow
Stephen C
SC
D000409:Alanine / Q000235:genetics; D020816:Amino Acid Motifs; D001057:Apolipoproteins E / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D002118:Calcium / Q000737:chemistry; D002151:Calorimetry; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry; D008024:Ligands; D026503:Low Density Lipoprotein Receptor-Related Protein-1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008239:Lysine / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D011233:Precipitin Tests; D011485:Protein Binding / Q000235:genetics; D017510:Protein Folding; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000235:genetics; D011973:Receptors, LDL / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid
D001057:Apolipoproteins E; D004220:Disulfides; D008024:Ligands; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D020033:Protein Isoforms; D011973:Receptors, LDL; D008239:Lysine; D000409:Alanine; D002118:Calcium; D004134:Dimyristoylphosphatidylcholine