title: Reconstruction of LPS Transfer Cascade Reveals Structural Determinants within LBP, CD14, and TLR4-MD2 for Efficient LPS Recognition and Transfer.
abstract: Lipopolysaccharide (LPS), the major component of the outer membrane of Gram-negative bacteria, binds Toll-like receptor 4 (TLR4)-MD2 complex and activates innate immune responses. LPS transfer to TLR4-MD2 is catalyzed by both LPS binding protein (LBP) and CD14. To define the sequential molecular interactions underlying this transfer, we reconstituted in vitro the entire LPS transfer process from LPS micelles to TLR4-MD2. Using electron microscopy and single-molecule approaches, we characterized the dynamic intermediate complexes for LPS transfer: LBP-LPS micelles, CD14-LBP-LPS micelle, and CD14-LPS-TLR4-MD2 complex. A single LBP molecule bound longitudinally to LPS micelles catalyzed multi-rounds of LPS transfer to CD14s that rapidly dissociated from LPB-LPS complex upon LPS transfer via electrostatic interactions. Subsequently, the single LPS molecule bound to CD14 was transferred to TLR4-MD2 in a TLR4-dependent manner. The definition of the structural determinants of the LPS transfer cascade to TLR4 may enable the development of targeted therapeutics for intervention in LPS-induced sepsis.
authors.affiliation: National Creative Research Initiative Center for Single-Molecule Systems Biology, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 34141, Korea; Department of Physics, KAIST, Daejeon 34141, Korea.
authors.forename: Je-Kyung
authors.initials: JK
authors.lastname: Ryu
authors.affiliation: Biomedical Science and Engineering Interdisciplinary Program, KAIST, Daejeon 34141, Korea.
authors.forename: Soo Jin
authors.initials: SJ
authors.lastname: Kim
authors.affiliation: National Creative Research Initiative Center for Single-Molecule Systems Biology, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 34141, Korea; Department of Physics, KAIST, Daejeon 34141, Korea; Center for Nanomedicine, Institute for Basic Science (IBS), Yonsei University, Seoul 03722, Korea; Yonsei-IBS Institute, Yonsei University, Seoul 03722, Korea.
authors.forename: Sang-Hyun
authors.initials: SH
authors.lastname: Rah
authors.affiliation: Biomedical Science and Engineering Interdisciplinary Program, KAIST, Daejeon 34141, Korea.
authors.forename: Ji In
authors.initials: JI
authors.lastname: Kang
authors.affiliation: Biomedical Science and Engineering Interdisciplinary Program, KAIST, Daejeon 34141, Korea.
authors.forename: Hi Eun
authors.initials: HE
authors.lastname: Jung
authors.affiliation: Graduate School of Medical Science & Engineering, KAIST, Daejeon 34141, Korea.
authors.forename: Dongsun
authors.initials: D
authors.lastname: Lee
authors.affiliation: Biomedical Science and Engineering Interdisciplinary Program, KAIST, Daejeon 34141, Korea; Graduate School of Medical Science & Engineering, KAIST, Daejeon 34141, Korea.
authors.forename: Heung Kyu
authors.initials: HK
authors.lastname: Lee
authors.affiliation: Department of Chemistry, KAIST, Daejeon 34141, Korea.
authors.forename: Jie-Oh
authors.initials: JO
authors.lastname: Lee
authors.affiliation: Department of Biomedical Laboratory Science, College of Health Science, Eulji University, Seongnam 461-713, Korea.
authors.forename: Beom Seok
authors.initials: BS
authors.lastname: Park
authors.affiliation: National Creative Research Initiative Center for Single-Molecule Systems Biology, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 34141, Korea; Department of Physics, KAIST, Daejeon 34141, Korea; Center for Nanomedicine, Institute for Basic Science (IBS), Yonsei University, Seoul 03722, Korea; Yonsei-IBS Institute, Yonsei University, Seoul 03722, Korea. Electronic address: tyyoon@yonsei.ac.kr.
authors.forename: Tae-Young
authors.initials: TY
authors.lastname: Yoon
authors.affiliation: Biomedical Science and Engineering Interdisciplinary Program, KAIST, Daejeon 34141, Korea; Graduate School of Medical Science & Engineering, KAIST, Daejeon 34141, Korea. Electronic address: hm_kim@kaist.ac.kr.
authors.forename: Ho Min
authors.initials: HM
authors.lastname: Kim
mesh_terms: D000209:Acute-Phase Proteins / Q000276:immunology*; D000818:Animals; D002352:Carrier Proteins / Q000276:immunology*; D006801:Humans; D018950:Lipopolysaccharide Receptors / Q000276:immunology*; D008070:Lipopolysaccharides / Q000276:immunology*; D051218:Lymphocyte Antigen 96 / Q000276:immunology*; D008562:Membrane Glycoproteins / Q000276:immunology*; D051379:Mice; D046529:Microscopy, Electron, Transmission; D015398:Signal Transduction / Q000276:immunology; D051197:Toll-Like Receptor 4 / Q000276:immunology*; D000099231:Lipopolysaccharide-Binding Protein
chemical_list: D000209:Acute-Phase Proteins; D002352:Carrier Proteins; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D051218:Lymphocyte Antigen 96; D008562:Membrane Glycoproteins; D051197:Toll-Like Receptor 4; D000099231:Lipopolysaccharide-Binding Protein
keywords: CD14; LBP; LPS recognition; LPS transfer; TLR4/MD2; innate immunity; negative-stain EM; single-molecule fluorescence analysis