Solution state NMR structure and dynamics of KpOmpA, a 210 residue transmembrane domain possessing a high potential for immunological applications |
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Authors: | Renault Marie Saurel Olivier Czaplicki Jerzy Demange Pascal Gervais Virginie Löhr Frank Réat Valérie Piotto Martial Milon Alain |
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Institution: | 1 Université de Toulouse, UPS; Institute of Pharmacology and Structural Biology - IPBS; 205 rte de Narbonne, F-31077, Toulouse, France 2 CNRS; UMR 5089 - IPBS, Toulouse, France 3 University of Frankfurt, Center for Biomolecular Magnetic Resonance, Max von Laue-Str. 9, D-60438 Frankfurt am Main, Germany 4 BRUKER Biospin France, NMR Application Laboratory, 34, rue de l'industrie, F-67166 Wissembourg, France |
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Abstract: | The three-dimensional structure of the outer membrane protein A from Klebsiella pneumoniae transmembrane domain was determined by NMR. This protein induces specific humoral and cytotoxic responses, and is a potent carrier protein. This is one of the largest integral membrane proteins (210 residues) for which nearly complete resonance assignment, including side chains, has been achieved so far. The methodology rested on the use of 900 MHz 3D and 4D TROSY experiments recorded on a uniformly 15N,13C,2H-labeled sample and on a perdeuterated methyl protonated sample. The structure was refined from 920 experimental constraints, giving an ensemble of 20 best structures with an r.m.s. deviation of 0.54 Å for the main chain atoms in the core eight-stranded β-barrel. The protein dynamics was assessed, in a residue-specific manner, by 1H-15N NOEs (pico- to nanosecond timescale), exchange broadening (millisecond to second) and 1H-2H chemical exchange (hour-weeks). |
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Keywords: | MP membrane protein ZW 3-14 n-tetradecyl-N N-dimethyl-3-ammonio-1-propanesulfonate C8E4 n-octyltetraoxyethylene DPC docecylphosphocholine DHPC 1 2-dihexanoyl-sn-glycero-3-phosphocholine TROSY transverse relaxation optimized spectroscopy HMQC heteronuclear multiple quantum coherence NOESY nuclear Overhauser effect spectroscopy NOE nuclear Overhauser effect |
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