Structure of the SecY channel during initiation of protein translocation

Many secretory proteins are targeted by signal sequences to a protein-conducting channel, formed by prokaryotic SecY- or eukaryotic Sec61-complexes, and are translocated across the membrane during their synthesis1,2. Crystal structures of the inactive channel show that the SecY subunit of the heterotrimeric complex consists of two halves that form an hourglass-shaped pore with a constriction in the middle of the membrane and a lateral gate that faces the lipid phase3-5. The closed channel has an empty cytoplasmic funnel and an extracellular funnel that is filled with a small helical domain, called the plug. During initiation of translocation, a ribosome–nascent chain complex binds to the SecY/Sec61 complex, resulting in insertion of the nascent chain. However, the mechanism of channel opening during translocation is unclear. Here, we have addressed this question by determining structures of inactive and active ribosome–channel complexes with cryo-electron microscopy. Non-translating ribosome–SecY channel complexes derived from Methanococcus jannaschii or Escherichia coli show the channel in its closed state, and indicate that ribosome binding per se causes only minor changes. The structure of an active E. coli ribosome–channel complex demonstrates that the nascent chain opens the channel, causing mostly rigid body movements of the N- and C-terminal halves of SecY. In this early translocation intermediate, the polypeptide inserts as a loop into the SecY channel with the hydrophobic signal sequence intercalated into the open lateral gate. The nascent chain also forms a loop on the cytoplasmic surface of SecY rather than directly entering the channel

Unfixed cryosections of striated muscle to study dynamic molecular events.

The structures of the actin and myosin filaments of striated muscle have been studied extensively in the past by sectioning of fixed specimens. However, chemical fixation alters molecular details and prevents biochemically induced structural changes. To overcome these problems, we investigate here the potential of cryosectioning unfixed muscle. In cryosections of relaxed, unfixed specimens, individual myosin filaments displayed the characteristic helical organization of detached cross-bridges, but the filament lattice had disintegrated. To preserve both the filament lattice and the molecular structure of the filaments, we decided to section unfixed rigor muscle, stabilized by actomyosin cross-bridges. The best sections showed periodic, angled cross-bridges attached to actin and their Fourier transforms displayed layer lines similar to those in x-ray diffraction patterns of rigor muscle. To preserve relaxed filaments in their original lattice, unfixed sections of rigor muscle were picked up on a grid and relaxed before negative staining. The myosin and actin filaments showed the characteristic helical arrangements of detached cross-bridges and actin subunits, and Fourier transforms were similar to x-ray patterns of relaxed muscle. We conclude that the rigor structure of muscle and the ability of the filament lattice to undergo the rigor-relaxed transformation can be preserved in unfixed cryosections. In the future, it should be possible to carry out dynamic studies of active sacromeres by cryo-electron microscopy

Erratum: Structure of the 70S ribosome from human pathogen Staphylococcus aureus (Nucleic Acids Research (2017) DOI: 10.1093/nar/gkw933)

© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.The authors wish to correct their Funding statement as follows: FUNDING. 'Centre National de la Recherche Scientifique' (CNRS) and the 'Agence Nationale de la Recherche' as part of the 'Investissements d'Avenir' program [LabEx: ANR-10-LABX-0036-NETRNA to P.R., Y.H.; ANR-15-CE11-0021-01 to G.Y.]; 'Fondation pour la Recherche Médicale en France' [FDT20140930867 to I.K; 'European Research Council advanced grant' [294312 to M.Y.]; the 'Russian Science Foundation' [Project No. 16-14-10014 to I.K., M.Y.]. Funding for open access charge: Centre National de la Recherche Scientifique (CNRS). In addition, Marat Yusupov is associated with both affiliations1 and 2. The authors apologise to the readers for this error

Erratum: Structure of the 70S ribosome from human pathogen Staphylococcus aureus (Nucleic Acids Research (2017) DOI: 10.1093/nar/gkw933)

© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.The authors wish to correct their Funding statement as follows: FUNDING. 'Centre National de la Recherche Scientifique' (CNRS) and the 'Agence Nationale de la Recherche' as part of the 'Investissements d'Avenir' program [LabEx: ANR-10-LABX-0036-NETRNA to P.R., Y.H.; ANR-15-CE11-0021-01 to G.Y.]; 'Fondation pour la Recherche Médicale en France' [FDT20140930867 to I.K; 'European Research Council advanced grant' [294312 to M.Y.]; the 'Russian Science Foundation' [Project No. 16-14-10014 to I.K., M.Y.]. Funding for open access charge: Centre National de la Recherche Scientifique (CNRS). In addition, Marat Yusupov is associated with both affiliations1 and 2. The authors apologise to the readers for this error

Erratum: Structure of the 70S ribosome from human pathogen Staphylococcus aureus (Nucleic Acids Research (2017) DOI: 10.1093/nar/gkw933)

© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.The authors wish to correct their Funding statement as follows: FUNDING. 'Centre National de la Recherche Scientifique' (CNRS) and the 'Agence Nationale de la Recherche' as part of the 'Investissements d'Avenir' program [LabEx: ANR-10-LABX-0036-NETRNA to P.R., Y.H.; ANR-15-CE11-0021-01 to G.Y.]; 'Fondation pour la Recherche Médicale en France' [FDT20140930867 to I.K; 'European Research Council advanced grant' [294312 to M.Y.]; the 'Russian Science Foundation' [Project No. 16-14-10014 to I.K., M.Y.]. Funding for open access charge: Centre National de la Recherche Scientifique (CNRS). In addition, Marat Yusupov is associated with both affiliations1 and 2. The authors apologise to the readers for this error

Structure of the mammalian 80S ribosome at 87 angstrom resolution

In this paper, we present a structure of the mammalian ribosome determined at not, vert, similar8.7 Å resolution by electron cryomicroscopy and single-particle methods. A model of the ribosome was created by docking homology models of subunit rRNAs and conserved proteins into the density map. We then modeled expansion segments in the subunit rRNAs and found unclaimed density for not, vert, similar20 proteins. In general, many conserved proteins and novel proteins interact with expansion segments to form an integrated framework that may stabilize the mature ribosome. Our structure provides a snapshot of the mammalian ribosome at the beginning of translation and lends support to current models in which large movements of the small subunit and L1 stalk occur during tRNA translocation. Finally, details are presented for intersubunit bridges that are specific to the eukaryotic ribosome. We suggest that these bridges may help reset the conformation of the ribosome to prepare for the next cycle of chain elongation