Molecular chaperone RAP interacts with LRP1 in a dynamic bivalent mode and enhances folding of ligand-binding regions of other LDLR family receptors
Hernandez, Nancy E
Kurasawa, James H
Hassink, Gabriela U
Shestopal, Svetlana A
Strickland, Dudley K
Sarafanov, Andrey G
JournalJournal of Biological Chemistry
MetadataShow full item record
AbstractThe low-density lipoprotein receptor (LDLR) family of receptors are cell-surface receptors that internalize numerous ligands and play crucial role in various processes, such as lipoprotein metabolism, hemostasis, fetal development, etc. Previously, receptor-associated protein (RAP) was described as a molecular chaperone for LDLR-related protein 1 (LRP1), a prominent member of the LDLR family. We aimed to verify this role of RAP for LRP1 and two other LDLR family receptors, LDLR and vLDLR, and to investigate the mechanisms of respective interactions using a cell culture model system, purified system, and in silico modelling. Upon co-expression of RAP with clusters of the ligand-binding complement repeats (CRs) of the receptors in secreted form in insect cells culture, the isolated proteins had increased yield, enhanced folding, and improved binding properties compared to proteins expressed without RAP, as determined by circular dichroism and surface plasmon resonance. Within LRP1 CR-clusters II and IV, we identified multiple sites comprised of adjacent CR-doublets, which provide alternative bivalent binding combinations with specific pairs of lysines on RAP. Mutational analysis of these lysines within each of isolated RAP D1/D2 and D3 domains having high-affinity to LRP1, and of conserved tryptophans on selected CR-doublets of LRP1, as well as in silico docking of a model LRP1 CR-triplet with RAP indicated a universal role for these residues in interaction of RAP and LRP1. Consequently, we propose a new model of RAP interaction with LDLR family receptors based on switching of the bivalent contacts between molecules over time in a dynamic mode.
Rights/TermsCopyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.
KeywordLDL-Receptor Related Protein-Associated Protein (RAP)
factor VIII (FVIII)
low‐density lipoprotein (LDL)
Identifier to cite or link to this itemhttp://hdl.handle.net/10713/15895
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