title: NMR studies of hexaacylated endotoxin bound to wild-type and F126A mutant MD-2 and MD-2·TLR4 ectodomain complexes.
abstract: Host response to invasion by many gram-negative bacteria depends upon activation of Toll-like receptor 4 (TLR4) by endotoxin presented as a monomer bound to myeloid differentiation factor 2 (MD-2). Metabolic labeling of hexaacylated endotoxin (LOS) from Neisseria meningitidis with [(13)C]acetate allowed the use of NMR to examine structural properties of the fatty acyl chains of LOS present in TLR4-agonistic and -antagonistic binary and ternary complexes with, respectively, wild-type or mutant (F126A) MD-2 ± TLR4 ectodomain. Chemical shift perturbation indicates that Phe(126) affects the environment and/or position of each of the bound fatty acyl chains both in the binary LOS·MD-2 complex and in the ternary LOS·MD-2·TLR4 ectodomain complex. In both wild-type and mutant LOS·MD-2 complexes, one of the six fatty acyl chains of LOS is more susceptible to paramagnetic attenuation, suggesting protrusion of that fatty acyl chain from the hydrophobic pocket of MD-2, independent of association with TLR4. These findings indicate that re-orientation of the aromatic side chain of Phe(126) is induced by binding of hexaacylated E, preceding interaction with TLR4. This re-arrangement of Phe(126) may act as a "hydrophobic switch," driving agonist-dependent contacts needed for TLR4 dimerization and activation.
authors.affiliation: NMR Core Facility, Roy A. and Lucille J. Carver College of Medicine, University of Iowa, Iowa City, Iowa 52241, USA.
authors.forename: Liping
authors.initials: L
authors.lastname: Yu
authors.forename: Rachel L
authors.initials: RL
authors.lastname: Phillips
authors.forename: DeSheng
authors.initials: D
authors.lastname: Zhang
authors.forename: Athmane
authors.initials: A
authors.lastname: Teghanemt
authors.forename: Jerrold P
authors.initials: JP
authors.lastname: Weiss
authors.forename: Theresa L
authors.initials: TL
authors.lastname: Gioannini
mesh_terms: D000107:Acetylation; D019943:Amino Acid Substitution; D004731:Endotoxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D051218:Lymphocyte Antigen 96 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D009345:Neisseria meningitidis / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D051197:Toll-Like Receptor 4 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D004731:Endotoxins; C495377:LY96 protein, human; D051218:Lymphocyte Antigen 96; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4