Evidence That Factor VIII Forms a Bivalent Complex with the Low Density Lipoprotein (LDL) Receptor-related Protein 1 (LRP1)

Abstract

Hemophilia A is a bleeding disorder caused by a deficiency in coagulation factor VIII (fVIII) that affects 1 in 5,000 males. Current prophylactic replacement therapy, although effective, is difficult to maintain due to the cost and frequency of injections. Hepatic clearance of fVIII is mediated by the LDL receptor-related protein 1 (LRP1), a member of the LDL receptor family. Although it is well established that fVIII binds LRP1, the molecular details of this interaction are unclear as most of the studies have been performed using fragments of fVIII and LRP1. In the current investigation, we examine the binding of intact fVIII to full-length LRP1 to gain insight into the molecular interaction. Chemical modification studies confirm the requirement for lysine residues in the interaction of fVIII with LRP1. Examination of the ionic strength dependence of the interaction of fVIII with LRP1 resulted in a Debye-Hückel plot with a slope of 1.8 ± 0.5, suggesting the involvement of two critical charged residues in the interaction of fVIII with LRP1. Kinetic studies utilizing surface plasmon resonance techniques reveal that the high affinity of fVIII for LRP1 results from avidity effects mediated by the interactions of two sites in fVIII with complementary sites on LRP1 to form a bivalent fVIII·LRP1 complex. Furthermore, although fVIII bound avidly to soluble forms of clusters II and IV from LRP1, only soluble cluster IV competed with the binding of fVIII to full-length LRP1, revealing that cluster IV represents the major fVIII binding site in LRP1.

Complement C1q Interacts With LRP1 Clusters II and IV Through a Site Close but Different From the Binding Site of Its C1r and C1s-Associated Proteases

“…This might be difficult if we consider that the potential residues for LRP1 interaction are oriented towards the center of the C1q cone ( Figure 11). Some LRP1 ligands have been described to interact better with cluster IV than with cluster II which is also what we observe in our study (33,44). From that observation, one could hypothesize that the LRP1 ligands bind first to the cluster that is more distant from the membrane (cluster II) and then interact with cluster IV.…”

“…Moreover, the affinity of the C1r2s2 interaction with C1q is stronger than the one of soluble LRP1 clusters which is in favor of the C1 formation in this experimental setting [ A common feature of most of LDL-receptors ligands is their ability to bind to heparin, suggesting the implication of one or more highly positively charged regions in the recognition of the receptor. Indeed, a "Lysine ligand mode" of interaction with tandem CR modules has been described for the binding of LRP1 with its ligands, such as RAP (30), a2Macroglobulin (31), ApoE (32), and factor VIII (33). These interactions are calcium dependent and salt sensitive.…”

Recombinant factor VIII Fc fusion protein drives regulatory macrophage polarization

“…Receptors implicated in FVIII uptake are also present on monocytes/macrophages. 29 For example, LRP1/CD91, an endocytic receptor, has been shown to bind FVIII 30 and was found to be upregulated in monocytes from HemA patients. 31 As rFVIIIFc is administered IV, blood monocytes of HemA patients are the first innate immune cells to interact with the administered FVIII.…”

The low density receptor-related protein 1 plays a significant role in ricin-mediated intoxication of lung cells

“…In this study we identified cluster II as the sole LRP1 binding site for ricin, and demonstrated that this binding is mediated by subunit B of the toxin. Cluster II and IV, which are responsible for the majority of ligand binding to the LRP1 receptor 40,[57][58][59] are highly similar in their binding properties, displaying only minor differences regarding their kinetics of interactions 60 . Huang et al 61 , suggested that the CR modules within these clusters, present different charge densities and hydrophobic patches, which in turn lead to varying receptor-ligands interactions responsible for the different ligand specificity of each cluster.…”