Decoding of Lipoprotein−Receptor Interactions: Properties of Ligand Binding Modules Governing Interactions with Apolipoprotein E

Abstract

Clusters of complement-type ligand binding repeats in the LDL receptor family are thought to mediate the interactions between these receptors and their various ligands. Apolipoprotein E, a key ligand for cholesterol homeostasis, has been shown to interact with LDLR, LRP and VLDLR, through these clusters. LDLR and VLDLR each contain a single ligand-binding repeat cluster, whereas LRP contains three large clusters of ligand binding repeats, each with ligand binding functions. We show that within sLRP3, the three-repeat subcluster CR16-18 recapitulated ligand binding to the isolated receptor binding portion of . Binding experiments with LA3-5 of LDLR and CR16-18 showed that a conserved W25/D30 pair appears critical for high affinity binding to . The triple repeat LA3-5 showed the expected interaction with ApoE(1-191)•DMPC, but surprisingly CR16-18 did not interact with this form of ApoE. To understand these differences in ApoE binding affinity, we introduced mutations of conserved residues from LA5 into CR18, and produced a CR16-18 variant capable of binding •DMPC. This change cannot fully be accounted for by the interaction with the proposed ApoE receptor binding region, therefore we speculate that LA5 is recognizing a distinct epitope on ApoE that may only exists in the lipid bound form. The combination of avidity effects with this distinct recognition process likely governs the ApoE-LDL receptor interaction.The low density lipoprotein (LDL) superfamily of receptors mediates cholesterol uptake into cells (1). Members of this family share many structural characteristics and sequence homology including an extracellular ligand binding domain consisting of complement-type repeats (CRs), also called ligand binding modules (LAs), epidermal growth factor precursor homology repeats (EGFs), β-propeller domains, and a single transmembrane segment with an intracellular domain (Fig. 1). The most well characterized of these receptors, LDLR, is genetically linked to hypercholesterolemia (2). LDLR family members recognize apolipoproteins on the surface of lipid particles, and Apolipoprotein E, in particular, plays an important role in receptor mediated cholesterol uptake (3). Although LDLR is the primary receptor for cholesterol carrying lipoproteins, studies have shown that the LDL receptor-related protein (LRP) and the very low density lipoprotein receptor (VLDL) also mediate the uptake of ApoE enriched β-VLDLs (4-7).*To whom correspondence should be addressed: ph: (858) FAX: (858) 534-6174, ekomives@ucsd.edu. This supplemental material is available free of charge via the Internet at http://pubs.acs.org.Supporting Information: Supporting information is provided with this manuscript that includes: additional surface plasmon resonance data supporting the binding of CR16-18 to ApoE(141-155) 2 (Suppl. Figure 1); HSQC spectra showing the shifts in amide resonances during NMR titration experiments of CR16, CR17, CR18, LA3, LA4, and LA5 with Ub-ApoE(130-149) (Suppl. Figure 2); HSQC spectra showing the shifts in amide reso...

Initiating protease with modular domains interacts with β-glucan recognition protein to trigger innate immune response in insects

“…Analysis of the binding isotherm with an N-identical binding sites model showed that rHP14-LA bound calcium with an apparent affinity of K D ∼ 3 μM, and thermodynamic parameters of ΔH = −18 kcal/mol, and ΔS = −34.3 cal/mol deg. These values are comparable to those obtained for LAs (CR17 and 18) from the human LDLR family (20). The calcium concentration in hemolymph is ∼5 mM (21) and, thus, likely sufficient to saturate HP14 LAs.…”

Cluster III of Low-Density Lipoprotein Receptor-Related Protein 1 Binds Activated Blood Coagulation Factor VIII

“…Indeed, the cluster's binding site for ApoE is presented by a CR triplet (CR.16-18), and the strongest binding CR doublet (CR.17-18) to a relevant ApoE peptide was 10-fold weaker than with the triplet. 47 At the same time, the RAP binding can be relevant to both models. In support for the first model, such binding was proposed to involve simultaneously up to six adjacent CRs of a receptor 40 .…”

SorLA Complement-type Repeat Domains Protect the Amyloid Precursor Protein against Processing

“…The ligands bind to the receptor using side chains from a lysine and a hydrophobic residue (37)(38)(39)(40). This simple motif has been identified in many complexes between ligands and CR-domains, including a minor group of human rhinovirus and three CR-domains from the very low density lipoprotein receptor (41), Reelin, in complex with two CR-domains from the apolipoprotein E receptor 2 (42, 43), ␤2-microglobulin in complex with the fourth CR-domain from LDLR (44,45), and apolipoprotein E with CR17 from LRP1 (46,47). These protein complexes represent classical extracellular ligands binding to endocytosis receptors; dissociation of the complex in the acidic endosome allows degradation of the ligands upon further transport to the lysosome and recycling of the receptors to the cell surface (48).…”