title: Local and Global Dynamics in Klebsiella pneumoniae Outer Membrane Protein a in Lipid Bilayers Probed at Atomic Resolution.
abstract: The role of membrane proteins in cellular mechanism strongly depends on their dynamics, and solid-state magic-angle spinning (MAS) nuclear magnetic resonance (NMR) is a unique method to exhaustively characterize motions of proteins in a lipid environment. Herein, we make use of advances in1H-detected MAS NMR to describe the dynamics of the membrane domain of the Outer membrane protein A of Klebsiella pneumoniae (KpOmpA). By measuring1H-15N dipolar-coupling as well as15N R1and R1ρrelaxation rates at fast (60 kHz) MAS and high magnetic field (1 GHz), we were able to describe the motions of the residues of the β-barrel as a collective rocking of low amplitude and of hundreds of nanoseconds time scale. Residual local motions at the edges of the strands, underscored by enhanced15N R1ρrelaxation rates, report on the mobility of the connected loops. In agreement with MAS NMR data, proteolysis experiments performed on the full length KpOmpA as well as on its membrane domain, reconstituted in liposomes or in detergent micelles, revealed in all cases the existence of a unique trypsin cleavage site within the membrane domain (out of 16 potential Lys and Arg sites). This site is located in the extracellular loop L3, showing that it is highly accessible to protein-protein interactions. KpOmpA is involved in cell-cell recognition, for adhesion and immune response mechanisms. The L3 region may therefore play a key role in pathogenicity.
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale (CNRS/Université Paul Sabatier), Université de Toulouse, 31077 Toulouse, France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Saurel
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale (CNRS/Université Paul Sabatier), Université de Toulouse, 31077 Toulouse, France.
authors.forename: Iordan
authors.initials: I
authors.lastname: Iordanov
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale (CNRS/Université Paul Sabatier), Université de Toulouse, 31077 Toulouse, France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Nars
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale (CNRS/Université Paul Sabatier), Université de Toulouse, 31077 Toulouse, France.
authors.forename: Pascal
authors.initials: P
authors.lastname: Demange
authors.affiliation: Institut de Sciences Analytiques (UMR 5280 CNRS/ENS-Lyon/UCB Lyon 1), Université de Lyon, 69007 Lyon, France.
authors.forename: Tanguy
authors.initials: T
authors.lastname: Le Marchand
authors.affiliation: Institut de Sciences Analytiques (UMR 5280 CNRS/ENS-Lyon/UCB Lyon 1), Université de Lyon, 69007 Lyon, France.
authors.forename: Loren B
authors.initials: LB
authors.lastname: Andreas
authors.affiliation: Institut de Sciences Analytiques (UMR 5280 CNRS/ENS-Lyon/UCB Lyon 1), Université de Lyon, 69007 Lyon, France.
authors.forename: Guido
authors.initials: G
authors.lastname: Pintacuda
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale (CNRS/Université Paul Sabatier), Université de Toulouse, 31077 Toulouse, France.
authors.forename: Alain
authors.identifier: 0000-0001-5692-8201
authors.initials: A
authors.lastname: Milon
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D007711:Klebsiella pneumoniae / Q000737:chemistry* / Q000378:metabolism; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D013058:Mass Spectrometry; D019906:Nuclear Magnetic Resonance, Biomolecular; D059748:Proteolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D013816:Thermodynamics
chemical_list: D001425:Bacterial Outer Membrane Proteins; D008051:Lipid Bilayers; D011994:Recombinant Proteins; C078167:OMPA outer membrane proteins