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Structural dynamics of the magnesium-bound conformation of CorA in a lipid bilayer.

ABSTRACT: The transmembrane conformation of Thermotoga maritima CorA, a magnesium transport system, has been studied in its Mg(2+)-bound form by site-directed spin labeling and electron paramagnetic resonance spectroscopy. Probe mobility together with accessibility data were used to evaluate the overall dynamics and relative arrangement of individual transmembrane segments TM1 and TM2. TM1 extends toward the cytoplasmic side creating a water-filled cavity, while TM2 is located in the periphery of the oligomer, contacting the lipid bilayer. A structural model for the conserved extracellular loop was generated based on EPR data and MD simulations, in which residue E316 is located toward the five-fold symmetry axis in position to electrostatically influence divalent ion translocation. Electrostatic analysis of our model suggest that, in agreement with the crystal structure, Mg(2+) -bound CorA is in a closed conformation. The present results suggest that long-range structural rearrangements are necessary to allow Mg(2+) translocation.

SUBMITTER: Dalmas O

PROVIDER: S-EPMC2935691 | biostudies-literature | 2010 Jul

REPOSITORIES: biostudies-literature

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Structural dynamics of the magnesium-bound conformation of CorA in a lipid bilayer.

Dalmas Olivier O Cuello Luis G LG Jogini Vishwanath V Cortes D Marien DM Roux Benoit B Perozo Eduardo E

Structure (London, England : 1993) 20100701 7

The transmembrane conformation of Thermotoga maritima CorA, a magnesium transport system, has been studied in its Mg(2+)-bound form by site-directed spin labeling and electron paramagnetic resonance spectroscopy. Probe mobility together with accessibility data were used to evaluate the overall dynamics and relative arrangement of individual transmembrane segments TM1 and TM2. TM1 extends toward the cytoplasmic side creating a water-filled cavity, while TM2 is located in the periphery of the olig ...[more]

PMID: 20637423

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Project description:The human voltage dependent anion channel 1 (VDAC) is a 32 kDa ?-barrel integral membrane protein that controls the transport of ions across the outer mitochondrial membrane. Despite the determination of VDAC solution and diffraction structures, a structural basis for the mechanism of its function is not yet fully understood. Biophysical studies suggest VDAC requires a lipid bilayer to achieve full function, motivating the need for atomic resolution structural information of VDAC in a membrane environment. Here we report an essential step toward that goal: extensive assignments of backbone and side chain resonances for VDAC in DMPC lipid bilayers via magic angle spinning nuclear magnetic resonance (MAS NMR). VDAC reconstituted into DMPC lipid bilayers spontaneously forms two-dimensional lipid crystals, showing remarkable spectral resolution (0.5-0.3 ppm for (13)C line widths and <0.5 ppm (15)N line widths at 750 MHz). In addition to the benefits of working in a lipid bilayer, several distinct advantages are observed with the lipid crystalline preparation. First, the strong signals and sharp line widths facilitated extensive NMR resonance assignments for an integral membrane ?-barrel protein in lipid bilayers by MAS NMR. Second, a large number of residues in loop regions were readily observed and assigned, which can be challenging in detergent-solubilized membrane proteins where loop regions are often not detected due to line broadening from conformational exchange. Third, complete backbone and side chain chemical shift assignments could be obtained for the first 25 residues, which comprise the functionally important N-terminus. The reported assignments allow us to compare predicted torsion angles for VDAC prepared in DMPC 2D lipid crystals, DMPC liposomes, and LDAO-solubilized samples to address the possible effects of the membrane mimetic environment on the conformation of the protein. Concluding, we discuss the strengths and weaknesses of the reported assignment approach and the great potential for even more complete assignment studies and de novo structure determination via (1)H detected MAS NMR.

Dataset Information

Structural dynamics of the magnesium-bound conformation of CorA in a lipid bilayer.

Publications

Structural dynamics of the magnesium-bound conformation of CorA in a lipid bilayer.

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