title: Dynamics and binding affinity of spin-labeled stearic acids in β-lactoglobulin: evidences from EPR spectroscopy and molecular dynamics simulation.
abstract: β-Lactoglobulin (β-LG) is a member of the lipocalin protein family involved in the transport of fatty acids and other small hydrophobic molecules. The main binding site is at a central cavity, referred to as "calyx", formed by the protein β-barrel sandwich. Continuous-wave and pulsed Fourier transform electron spin resonance (cw- and FT-EPR) spectroscopy and molecular dynamics (MD) simulation were combined to investigate the interaction of fatty acids with bovine β-LG. Stearic acid bearing the nitroxide label at different positions, n, along the acyl chain (n-SASL, n = 5, 7, 10, 12, 16) were used. The EPR data show that the protein affinity for SASL decreases on going from n = 5 to 16. This behavior is due to the accommodation of the SASL in the protein calyx, which is hampered by steric hindrance of the doxyl ring for n ≥ 10, as evidenced by MD data. Conformation and dynamics of 5-SASL are similar to those of the unlabeled stearate molecule. 5-SASL in the protein binding site undergoes librational motion of small amplitude on the nanosecond time scale at cryogenic temperature and rotational dynamics with correlation time of 4.2 ns at physiological temperature. The results highlight the dynamical features of fatty acids/β-LG interaction.
authors.affiliation: Department of Physics, University of Calabria, Ponte P. Bucci 31C, 87036 Rende (CS), Italy. rita.guzzi@fis.unical.it
authors.forename: Rita
authors.initials: R
authors.lastname: Guzzi
authors.forename: Bruno
authors.initials: B
authors.lastname: Rizzuti
authors.forename: Rosa
authors.initials: R
authors.lastname: Bartucci
mesh_terms: D000818:Animals; D001665:Binding Sites; D002417:Cattle; D004578:Electron Spin Resonance Spectroscopy; D007782:Lactoglobulins / Q000737:chemistry*; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D013113:Spin Labels; D013229:Stearic Acids / Q000737:chemistry*
chemical_list: D007782:Lactoglobulins; D013113:Spin Labels; D013229:Stearic Acids