title: NMR investigation of the equilibrium partitioning of a water-soluble bile salt protein carrier to phospholipid vesicles.
abstract: Membrane binding by cytosolic fatty acid binding proteins (FABP) appears to constitute a key step of intracellular lipid trafficking. We applied NMR spectroscopy to study the partitioning of a water-soluble bile acid binding protein (BABP), belonging to the FABP family, between its free and lipid-vesicle-bound states. As the lipid-bound protein was NMR-invisible, the signals of the free biomolecule were analyzed to obtain quantitative information on binding affinity and steady-state kinetics. The data indicated a reversible interaction of BABP with anionic vesicles occurring in a very slow exchange regime on the NMR time scale. The approximate binding epitope was demonstrated from results on BABP samples in which different positively charged lysine residues were mutated to neutral alanines. H/D exchange measurements indicated a higher exposure to solvent for the core amino acid residues in the liposome-bound state. Finally, the BABP-liposome interaction was also investigated for the first time through an MRI-chemical exchange saturation transfer experiment that has potential applications not only in the field of biology, but also in biomedicine, bioanalytical chemistry, and nanotechnology.
authors.affiliation: Department of Biotechnology, University of Verona, 37134, Verona, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Ceccon
authors.forename: Mariapina
authors.initials: M
authors.lastname: D'Onofrio
authors.forename: Serena
authors.initials: S
authors.lastname: Zanzoni
authors.forename: Dario Livio
authors.initials: DL
authors.lastname: Longo
authors.forename: Silvio
authors.initials: S
authors.lastname: Aime
authors.forename: Henriette
authors.initials: H
authors.lastname: Molinari
authors.forename: Michael
authors.initials: M
authors.lastname: Assfalg
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D002645:Chickens; D041961:Deuterium Exchange Measurement; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D009994:Osmolar Concentration; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012995:Solubility
chemical_list: D002352:Carrier Proteins; D008081:Liposomes; D008562:Membrane Glycoproteins; D010743:Phospholipids; D011994:Recombinant Proteins; C037643:bile acid binding proteins
keywords: NMR spectroscopy; amphitropism; bile acid binding protein; chemical exchange saturation transfer; lipid vesicle