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81.
Complete structure of p97/valosin-containing protein reveals communication between nucleotide domains 总被引:2,自引:0,他引:2
The ATPase p97/VCP affects multiple events within the cell. These events include the alteration of both nuclear and mitotic Golgi membranes, the dislocation of ubiquitylated proteins from the endoplasmic reticulum and regulation of the NF-kappa b pathway. Here we present the crystal structure of full-length Mus musculus p97/VCP in complex with a mixture of ADP and ADP-AlF(x) at a resolution of 4.7 A. This is the first complete hexameric structure of a protein containing tandem AAA (ATPases associated with a variety of cellular activities) domains. Comparison of the crystal structure and cryo-electron microscopy (EM) reconstructions reveals large conformational changes in the helical subdomains during the hydrolysis cycle. Structural and functional data imply a communication mechanism between the AAA domains. A Zn(2+) occludes the central pore of the hexamer, suggesting that substrate does not thread through the pore of the molecule. 相似文献
82.
Brunger AT 《Structure (London, England : 1993)》2005,13(2):171-172
The threshold of what is considered "acceptable" resolution for obtaining mechanistic insights is being pushed by recent structures at 3.8 to 4.7 A resolution. One of these structures, that of a fully glycosylated SIV gp120 envelope glycoprotein in an unliganded conformation at 4.0 A resolution, is described in this issue (Chen et al., 2005). 相似文献
83.
MOTIVATION: Subcellular protein localization data are critical to the quantitative understanding of cellular function and regulation. Such data are acquired via observation and quantitative analysis of fluorescently labeled proteins in living cells. Differentiation of labeled protein from cellular artifacts remains an obstacle to accurate quantification. We have developed a novel hybrid machine-learning-based method to differentiate signal from artifact in membrane protein localization data by deriving positional information via surface fitting and combining this with fluorescence-intensity-based data to generate input for a support vector machine. RESULTS: We have employed this classifier to analyze signaling protein localization in T-cell activation. Our classifier displayed increased performance over previously available techniques, exhibiting both flexibility and adaptability: training on heterogeneous data yielded a general classifier with good overall performance; training on more specific data yielded an extremely high-performance specific classifier. We also demonstrate accurate automated learning utilizing additional experimental data. 相似文献
84.
Gottschalk KE Adams PD Brunger AT Kessler H 《Protein science : a publication of the Protein Society》2002,11(7):1800-1812
Integrins are composed of noncovalently bound dimers of an alpha- and a beta-subunit. They play an important role in cell-matrix adhesion and signal transduction through the cell membrane. Signal transduction can be initiated by the binding of intracellular proteins to the integrin. Binding leads to a major conformational change. The change is passed on to the extracellular domain through the membrane. The affinity of the extracellular domain to certain ligands increases; thus at least two states exist, a low-affinity and a high-affinity state. The conformations and conformational changes of the transmembrane (TM) domain are the focus of our interest. We show by a global search of helix-helix interactions that the TM section of the family of integrins are capable of adopting a structure similar to the structure of the homodimeric TM protein Glycophorin A. For the alpha(IIb)beta(3) integrin, this structural motif represents the high-affinity state. A second conformation of the TM domain of alpha(IIb)beta(3) is identified as the low-affinity state by known mutational and nuclear magnetic resonance (NMR) studies. A transition between these two states was determined by molecular dynamics (MD) calculations. On the basis of these calculations, we propose a three-state mechanism. 相似文献
85.
Structural basis of Rab effector specificity: crystal structure of the small G protein Rab3A complexed with the effector domain of rabphilin-3A 总被引:15,自引:0,他引:15
The small G protein Rab3A plays an important role in the regulation of neurotransmitter release. The crystal structure of activated Rab3A/GTP/Mg2+ bound to the effector domain of rabphilin-3A was solved to 2.6 A resolution. Rabphilin-3A contacts Rab3A in two distinct areas. The first interface involves the Rab3A switch I and switch II regions, which are sensitive to the nucleotide-binding state of Rab3A. The second interface consists of a deep pocket in Rab3A that interacts with a SGAWFF structural element of rabphilin-3A. Sequence and structure analysis, and biochemical data suggest that this pocket, or Rab complementarity-determining region (RabCDR), establishes a specific interaction between each Rab protein and its effectors. RabCDRs could be major determinants of effector specificity during vesicle trafficking and fusion. 相似文献
86.
Burnett JC Ruthel G Stegmann CM Panchal RG Nguyen TL Hermone AR Stafford RG Lane DJ Kenny TA McGrath CF Wipf P Stahl AM Schmidt JJ Gussio R Brunger AT Bavari S 《The Journal of biological chemistry》2007,282(7):5004-5014
An efficient research strategy integrating empirically guided, structure-based modeling and chemoinformatics was used to discover potent small molecule inhibitors of the botulinum neurotoxin serotype A light chain. First, a modeled binding mode for inhibitor 2-mercapto-3-phenylpropionyl-RATKML (K(i) = 330 nM) was generated, and required the use of a molecular dynamic conformer of the enzyme displaying the reorientation of surface loops bordering the substrate binding cleft. These flexible loops are conformationally variable in x-ray crystal structures, and the model predicted that they were pivotal for providing complementary binding surfaces and solvent shielding for the pseudo-peptide. The docked conformation of 2-mercapto-3-phenylpropionyl-RATKML was then used to refine our pharmacophore for botulinum serotype A light chain inhibition. Data base search queries derived from the pharmacophore were employed to mine small molecule (non-peptidic) inhibitors from the National Cancer Institute's Open Repository. Four of the inhibitors possess K(i) values ranging from 3.0 to 10.0 microM. Of these, NSC 240898 is a promising lead for therapeutic development, as it readily enters neurons, exhibits no neuronal toxicity, and elicits dose-dependent protection of synaptosomal-associated protein (of 25 kDa) in a primary culture of embryonic chicken neurons. Isothermal titration calorimetry showed that the interaction between NSC 240898 and the botulinum A light chain is largely entropy-driven, and occurs with a 1:1 stoichiometry and a dissociation constant of 4.6 microM. 相似文献
87.
88.
Sandro Vivona Corey W. Liu Pavel Strop Valeria Rossi Francesco Filippini Axel T. Brunger 《The Journal of biological chemistry》2010,285(23):17965-17973
SNARE protein complexes are key mediators of exocytosis by juxtaposing opposing membranes, leading to membrane fusion. SNAREs generally consist of one or two core domains that can form a four-helix bundle with other SNARE core domains. Some SNAREs, such as syntaxin target-SNAREs and longin vesicular-SNAREs, have independent, folded N-terminal domains that can interact with their respective SNARE core domains and thereby affect the kinetics of SNARE complex formation. This autoinhibition mechanism is believed to regulate the role of the longin VAMP7/TI-VAMP in neuronal morphogenesis. Here we use nuclear magnetic resonance spectroscopy to study the longin-SNARE core domain interaction for VAMP7. Using complete backbone resonance assignments, chemical shift perturbations analysis, and hydrogen/deuterium exchange experiments, we conclusively show that VAMP7 adopts a preferentially closed conformation in solution. Taken together, the closed conformation of longins is conserved, in contrast to the syntaxin family of SNAREs for which mixtures of open and closed states have been observed. This may indicate different regulatory mechanisms for SNARE complexes containing syntaxins and longins, respectively. 相似文献
89.
90.
Brunger AT 《Current opinion in neurobiology》2000,10(3):293-302
The fusion of vesicles with target membranes is controlled by a complex network of protein-protein and protein-lipid interactions. Recent structures of the SNARE complex, synaptotagmin III, nSec1, domains of NSF and its adaptor SNAP, along with Rab3 and some of its effectors, provide the framework for developing molecular models of vesicle fusion and for designing experiments to test these models. Ultimately, this knowledge of the structures of higher-order complexes and their dynamic behavior will allow us to obtain a full understanding of the vesicle fusion protein machinery. 相似文献