Journal article
Protein-lipid interactions with Fusobacterium nucleatum major outer membrane protein FomA: spin-label EPR and polarized infrared spectroscopy

Publication Details
Anbazhagan, V.; Vijay, N.; Kleinschmidt, J.; Marsh, D.
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FomA, the major outer membrane protein of Fusobacterium nucleatum, was expressed and purified in Escherichia coli and reconstituted from detergent in bilayer membranes of phosphatidylcholines with chain lengths from C(12:0) to C(17:0). The conformation and orientation of membrane-incorporated FomA were determined from polarized, attenuated total reflection, infrared (IR) spectroscopy, and lipid-protein interactions with FomA were characterized by using electron paramagnetic resonance (EPR) spectroscopy of spin-labeled lipids. Approximately 190 residues of membranous FomA are estimated to be in a beta-sheet configuration from IR band fitting, which is consistent with a 14-strand transmembrane beta-barrel structure. IR dichroism of FomA indicates that the beta-strands are tilted by approximately 45 degrees relative to the sheet/barrel axis and that the order parameter of the latter displays a discontinuity corresponding to hydrophobic matching with fluid C(13:0) lipid chains. The stoichiometry ( N b = 23 lipids/monomer) of lipid-protein interaction from EPR demonstrates that FomA is not trimeric in membranes of diC(14:0) phosphatidylcholine and is consistent with a monomeric beta-barrel of 14-16 strands. The pronounced selectivity of interaction found with anionic spin-labeled lipids places basic residues of the protein in the vicinity of the polar-apolar membrane interfaces, consistent with current topology models. Comparison with similar data from the 8- to 22-stranded E. coli outer membrane proteins, OmpA, OmpG, and FhuA, supports the above conclusions.

0 (Bacterial Outer Membrane Proteins), 0 (fomA protein, Fusobacterium), 0 (Membrane Lipids), 0 (Spin Labels), Bacterial Outer Membrane Proteins/analysis/metabolism, Electron Spin Resonance Spectroscopy, Fusobacterium nucleatum/metabolism, Membrane Lipids/analysis/metabolism, Protein Structure, Secondary, Spectrophotometry, Infrared, Spin Labels, Structure-Activity Relationship

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Last updated on 2019-25-07 at 17:25