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Dodecamerization and insertion of the outer membrane secretin PulD is entirely determined by the C-terminal half of the polypeptide (PulD-CS). In the absence of its cognate chaperone PulS, PulD-CS and PulD mislocalize to the inner membrane, from which they are extractable with detergents but not urea. Electron microscopy of PulD-CS purified from the inner(More)
The specialized type III secretion (T3S) apparatus of pathogenic and symbiotic Gram-negative bacteria comprises a complex transmembrane organelle and an ATPase homologous to the F1-ATPase beta subunit. The T3S ATPase HrcN of Pseudomonas syringae associates with the inner membrane, and its ATP hydrolytic activity is stimulated by dodecamerization. The(More)
The introduction of direct electron detectors (DED) to cryo-electron microscopy has tremendously increased the signal-to-noise ratio (SNR) and quality of the recorded images. We discuss the optimal use of DEDs for cryo-electron crystallography, introduce a new automatic image processing pipeline, and demonstrate the vast improvement in the resolution(More)
The vacuolar protein sorting (Vps) pathway, in which Vps4 class I AAA-ATPases play a central role, regulates growth factor receptors, immune response, and developmental signaling, and participates in tumor suppression, apoptosis, and retrovirus budding. We present the first atomic structure of the nucleotide-free yeast His(6)DeltaNVps4p dimer and its AMPPNP(More)
For cells to function properly, membrane proteins must be able to diffuse within biological membranes. The functions of these membrane proteins depend on their position and also on protein-protein and protein-lipid interactions. However, so far, it has not been possible to study simultaneously the structure and dynamics of biological membranes. Here, we(More)
BACKGROUND Rh glycoproteins (RhAG, RhBG, RhCG) are members of the Amt/Mep/Rh family which facilitate movement of ammonium across plasma membranes. Changes in ammonium transport activity following expression of Rh glycoproteins have been described in different heterologous systems such as yeasts, oocytes and eukaryotic cell lines. However, in these complex(More)
Cyclic nucleotide-modulated ion channels are important for signal transduction and pacemaking in eukaryotes. The molecular determinants of ligand gating in these channels are still unknown, mainly because of a lack of direct structural information. Here we report ligand-induced conformational changes in full-length MloK1, a cyclic nucleotide-modulated(More)
UNLABELLED In Gram-negative bacteria, the Lol and Bam machineries direct the targeting of lipidated and nonlipidated proteins, respectively, to the outer membrane (OM). Using Pseudomonas aeruginosa strains with depleted levels of specific Bam and Lol proteins, we demonstrated a variable dependence of different OM proteins on these targeting pathways.(More)
We have built and extensively tested a tool-chain to prepare and screen two-dimensional crystals of membrane proteins by transmission electron microscopy (TEM) at room temperature. This automated process is an extension of a new procedure described recently that allows membrane protein 2D crystallization in parallel (Iacovache et al., 2010). The system(More)
MuRF1 is an E3 ubiquitin ligase central to muscle catabolism. It belongs to the TRIM protein family characterized by a tripartite fold of RING, B-box and coiled-coil (CC) motifs, followed by variable C-terminal domains. The CC motif is hypothesized to be responsible for domain organization in the fold as well as for high-order assembly into functional(More)