title: Evidence for Two Distinct Binding Sites for Lipoprotein Lipase on Glycosylphosphatidylinositol-anchored High Density Lipoprotein-binding Protein 1 (GPIHBP1).
abstract: GPIHBP1 is an endothelial membrane protein that transports lipoprotein lipase (LPL) from the subendothelial space to the luminal side of the capillary endothelium. Here, we provide evidence that two regions of GPIHBP1, the acidic N-terminal domain and the central Ly6 domain, interact with LPL as two distinct binding sites. This conclusion is based on comparative binding studies performed with a peptide corresponding to the N-terminal domain of GPIHBP1, the Ly6 domain of GPIHBP1, wild type GPIHBP1, and the Ly6 domain mutant GPIHBP1 Q114P. Although LPL and the N-terminal domain formed a tight but short lived complex, characterized by fast on- and off-rates, the complex between LPL and the Ly6 domain formed more slowly and persisted for a longer time. Unlike the interaction of LPL with the Ly6 domain, the interaction of LPL with the N-terminal domain was significantly weakened by salt. The Q114P mutant bound LPL similarly to the N-terminal domain of GPIHBP1. Heparin dissociated LPL from the N-terminal domain, and partially from wild type GPIHBP1, but was unable to elute the enzyme from the Ly6 domain. When LPL was in complex with the acidic peptide corresponding to the N-terminal domain of GPIHBP1, the enzyme retained its affinity for the Ly6 domain. Furthermore, LPL that was bound to the N-terminal domain interacted with lipoproteins, whereas LPL bound to the Ly6 domain did not. In summary, our data suggest that the two domains of GPIHBP1 interact independently with LPL and that the functionality of LPL depends on its localization on GPIHBP1.
authors: From the Department of Chemistry, Tallinn University of Technology, Tallinn 12618, Estonia.
authors: Mart
authors: M
authors: Reimund
authors: the Department of Medical Biosciences, Umeå University, SE-901 87 Umeå, Sweden, and.
authors: Mikael
authors: M
authors: Larsson
authors: the Department of Medical Biosciences, Umeå University, SE-901 87 Umeå, Sweden, and.
authors: Oleg
authors: O
authors: Kovrov
authors: the Institute of Technology, University of Tartu, Tartu 50411, Estonia.
authors: Sergo
authors: S
authors: Kasvandik
authors: the Department of Medical Biosciences, Umeå University, SE-901 87 Umeå, Sweden, and.
authors: Gunilla
authors: G
authors: Olivecrona
authors: From the Department of Chemistry, Tallinn University of Technology, Tallinn 12618, Estonia, aivar.lookene@ttu.ee.
authors: Aivar
authors: A
authors: Lookene
mesh_terms: D000818:Animals; D016880:Anisotropy; D001665:Binding Sites; D002417:Cattle; D003432:Cross-Linking Reagents / Q000737:chemistry; D004730:Endothelium, Vascular / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D017261:Glycosylphosphatidylinositols / Q000737:chemistry*; D006493:Heparin / Q000737:chemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008071:Lipoprotein Lipase / Q000737:chemistry*; D008074:Lipoproteins / Q000737:chemistry*; D013058:Mass Spectrometry; D051379:Mice; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats; D018110:Receptors, Lipoprotein / Q000737:chemistry*; D020349:Surface Plasmon Resonance
chemical_list: D003432:Cross-Linking Reagents; D000939:Epitopes; D005456:Fluorescent Dyes; C472779:GPI-HBP1 protein, mouse; C523701:GPIHBP1 protein, human; D017261:Glycosylphosphatidylinositols; D008074:Lipoproteins; D010455:Peptides; D018110:Receptors, Lipoprotein; D006493:Heparin; D008071:Lipoprotein Lipase
keywords: GPIHBP1; kinetics; lipoprotein lipase; lipoprotein metabolism; protein cross-linking; protein-protein interaction; surface plasmon resonance (SPR)