%0 Journal Article %J Proc Natl Acad Sci U S A %D 2009 %T Ribosome-induced changes in elongation factor Tu conformation control GTP hydrolysis. %A Villa, Elizabeth %A Sengupta, Jayati %A Trabuco, Leonardo G %A LeBarron, Jamie %A Baxter, Bill %A Shaikh, Tanvir R %A Grassucci, Robert A %A Nissen, Poul %A Ehrenberg, Måns %A Schulten, Klaus %A Frank, Joachim %K Cryoelectron Microscopy %K Enzyme Activation %K Escherichia coli %K Escherichia coli Proteins %K Guanosine Triphosphate %K Histidine %K Hydrolysis %K Hydrophobic and Hydrophilic Interactions %K Models, Molecular %K Peptide Elongation Factor Tu %K Protein Structure, Secondary %K Ribosomal Proteins %K Ribosomes %K RNA, Transfer %K Signal Transduction %X

In translation, elongation factor Tu (EF-Tu) molecules deliver aminoacyl-tRNAs to the mRNA-programmed ribosome. The GTPase activity of EF-Tu is triggered by ribosome-induced conformational changes of the factor that play a pivotal role in the selection of the cognate aminoacyl-tRNAs. We present a 6.7-A cryo-electron microscopy map of the aminoacyl-tRNA x EF-Tu x GDP x kirromycin-bound Escherichia coli ribosome, together with an atomic model of the complex obtained through molecular dynamics flexible fitting. The model reveals the conformational changes in the conserved GTPase switch regions of EF-Tu that trigger hydrolysis of GTP, along with key interactions, including those between the sarcin-ricin loop and the P loop of EF-Tu, and between the effector loop of EF-Tu and a conserved region of the 16S rRNA. Our data suggest that GTP hydrolysis on EF-Tu is controlled through a hydrophobic gate mechanism.

%B Proc Natl Acad Sci U S A %V 106 %P 1063-8 %8 01/2009 %G eng %U http://www.ncbi.nlm.nih.gov/pubmed/19122150 %N 4 %R 10.1073/pnas.0811370106 %0 Journal Article %J Nat Protoc %D 2008 %T SPIDER image processing for single-particle reconstruction of biological macromolecules from electron micrographs. %A Shaikh, Tanvir R %A Gao, Haixiao %A Baxter, Bill %A Asturias, Francisco J %A Boisset, Nicolas %A Leith, ArDean %A Frank, Joachim %K Image Processing, Computer-Assisted %K Microscopy, Electron %K Models, Molecular %K Molecular Structure %K Software %K User-Computer Interface %X

This protocol describes the reconstruction of biological molecules from the electron micrographs of single particles. Computation here is performed using the image-processing software SPIDER and can be managed using a graphical user interface, termed the SPIDER Reconstruction Engine. Two approaches are described to obtain an initial reconstruction: random-conical tilt and common lines. Once an existing model is available, reference-based alignment can be used, a procedure that can be iterated. Also described is supervised classification, a method to look for homogeneous subsets when multiple known conformations of the molecule may coexist.

%B Nat Protoc %V 3 %P 1941-74 %8 10/2008 %G eng %U http://www.ncbi.nlm.nih.gov/pubmed/19180078 %N 12 %R 10.1038/nprot.2008.156