CIISB Research Highlights Archive

  • J. Biol. Chem. 2017

    J. Biol. Chem. 2017

    Structural changes in crystal structure of of AfGcHK sensor protein induced by sodium dithionite soaking (PDB ID 5OHF) and conformational changes revealed by HDX-MS. (A) Two protein chains of the heme domain of the sensor protein observed in crystal structure including alternative B (yellow) occurring with dithionite soaking. (B) Conformational changes revealed by HDX-MS after 60 min of deuteration of the full-length AfGcHK proteins visualized on the protein structure. Differences between the Fe(III)-OH- form (active) and inactive Fe(II) form are color coded: grey - no difference, red - higher and blue - lower levels of deuteration.


    The heme-based oxygen sensor histidine kinase AfGcHK is part of a two-component signal transduction system in bacteria. O2 binding to the Fe(II) heme complex of its N-terminal globin domain strongly stimulates autophosphorylation at His-183 in its C-terminal kinase domain. The 6-coordinate heme Fe(III)-OH- and -CN- complexes of AfGcHK are also active, but the 5-coordinate heme Fe(II) complex and the heme-free apo-form are inactive. The crystal structures of the isolated dimeric globin domains of the active Fe(III)-CN- and inactive 5-coordinate Fe(II) forms were determined, revealing striking structural differences on the heme-proximal side of the globin domain. Using hydrogen/deuterium exchange coupled with mass spectrometry (HDX–MS) the intramolecular signal transduction mechanisms was investigated in full length AfGcHK.  The results suggest that structural changes at the heme proximal side, the globin domain-dimerization interface, and the ATP-binding site are important in the signal transduction mechanism of AfGcHK. For the first time, the conformational changes associated with signal transduction were studied in a full-length globin-coupled oxygen sensor protein and linked to directly observed structural changes in the globin domain.

    Stranava, M.; Man, P.; Skálová, T.; Kolenko, P.; Blaha, J.; Fojtikova, V.; Martínek, V.; Dohnálek, J.; Lengalova, A.; Rosůlek, M.; Shimizu, T. & Martínková, M.: Coordination and redox state-dependent structural changes of the heme-based oxygen sensor AfGcHK associated with intraprotein signal transduction. J. Biol. Chem. first Published on November 1, 2017, doi: 10.1074/jbc.M117.817023jbc.M117.817023.

  • J. Am. Chem. Soc. 2017

    J. Am. Chem. Soc. 2017

    High-resolution structure of a stable G-hairpin calculated from NMR data (PDB ID: 5M1W). (A) Ten lowest-energy structures. Loop residues are colored orange and O4′ atoms are colored red. (B) Schematic representation of hairpin folding topology. Chain reversal arrangement of the backbone and the 3′-to-5′ stacking of the terminal residues are indicated by dark green and magenta arrows, respectively. Anti and syn guanines that form G:G base pairs are colored dark and light blue, respectively.

    Lukáš Trantírek Research Group


    The first atomic resolution structure of a stable G-hairpin formed by a natively occurring DNA sequence is reported. An 11-nt long G-rich DNA oligonucleotide, 5′-d(GTGTGGGTGTG)-3′, corresponding to the most abundant sequence motif in irregular telomeric DNA from Saccharomyces cerevisiae adopts a novel type of mixed parallel/antiparallel fold-back DNA structure, which is stabilized by dynamic G:G base pairs that transit between N1-carbonyl symmetric and N1-carbonyl, N7-amino base-pairing arrangements. The structure reveals previously unknown principles of the folding of G-rich oligonucleotides that could be applied to the prediction of natural and/or the design of artificial recognition DNA elements. The structure also demonstrates that the folding landscapes of short DNA single strands is much more complex than previously assumed. 

    Gajarsky, M.; Zivkovic, M. L.; Stadlbauer, P.; Pagano, B.; Fiala, R.; Amato, J.; Tomaska, L.; Sponer, J.; Plavec, J. &  Trantirek, L.: Structure of a Stable G-Hairpin JACS 139, 3591-3594, doi:10.1021/jacs.6b10786 (2017)

  • PNAS 2017

    PNAS 2017

    A model of RNA polymerase II bound to the transcription termination factor Rtt103. The structural model was created using integrative structural biology.

    Richard Štefl Research Group


    RNA polymerase II (RNAPII) not only transcribes protein coding genes and many noncoding RNA, but also coordinates transcription and RNA processing. This coordination is mediated by a long C-terminal domain (CTD) of the largest RNAPII subunit, which serves as a binding platform for many RNA/protein-binding factors involved in transcription regulation. In this work, we used a hybrid approach to visualize the architecture of the full-length CTD in complex with the transcription termination factor Rtt103. Specifically, we first solved the structures of the isolated subcomplexes at high resolution and then arranged them into the overall envelopes determined at low resolution by small-angle X-ray scattering. The reconstructed overall architecture of the Rtt103–CTD complex reveals how Rtt103 decorates the CTD platform.

    Jasnovidova, O.; Klumpler, T.; Kubicek, K.; Kalynych, S.; Plevka, P. & Stefl, R.: Structure and dynamics of the RNAPII CTDsome with Rtt103, PNAS 2017 114 (42) 11133-11138; doi:10.1073/pnas.1712450114

  • Nucleic Acids Res. 2017

    Nucleic Acids Res. 2017

    Cryo-EM structure of the spinach chloroplast ribosome reveals the location of plastid-specific ribosomal proteins and extensions.

    Daniel N. Wilson Research Group


    Ribosomes are the protein synthesizing machines of the cell. Recent advances in cryo-EM have led to the determination of structures from a variety of species, including bacterial 70S and eukaryotic 80S ribosomes as well as mitoribosomes from eukaryotic mitochondria, however, to date high resolution structures of plastid 70S ribosomes have been lacking. Here we present a cryo-EM structure of the spinach chloroplast 70S ribosome, with an average resolution of 5.4 Å for the small 30S subunit and 3.6 Å for the large 50S ribosomal subunit. The structure reveals the location of the plastid-specific ribosomal proteins (RPs) PSRP1, PSRP4, PSRP5 and PSRP6 as well as the numerous plastid-specific extensions of the RPs. We discover many features by which the plastid-specific extensions stabilize the ribosome via establishing additional interactions with surrounding ribosomal RNA and RPs. Moreover, we identify a large conglomerate of plastid-specific protein mass adjacent to the tunnel exit site that could facilitate interaction of the chloroplast ribosome with the thylakoid membrane and the protein-targeting machinery. Comparing the Escherichia coli 70S ribosome with that of the spinach chloroplast ribosome provides detailed insight into the co-evolution of RP and rRNA.

    Graf, M.; Arenz, S.; Huter, P.; Dönhöfer, A.; Nováček, J. & Wilson D. N.: Cryo-EM structure of the spinach chloroplast ribosome reveals the location of plastid-specific ribosomal proteins and extensions. Nucleic Acids Research 45, 2887-2896, doi:10.1093/nar/gkw1272 (2017).

  • PNAS 2017

    PNAS 2017

    The virus structure of deformed wing virus of honeybees determined by cryo-electron microscopy. (A) Surface of the virus is rainbow-colored according to its distance from the particle center. (B) Cartoon representation of structure of the P-domain that decorates deformed wing virus surface is rainbow-colored from residue 260 in blue to 416 in red. The background shows image of the deformed wing virus particles from electron microscope.

    Pavel Plevka Research Group


    Honey bee populations in Europe and North America have been decreasing since the 1950s. Deformed wing virus (DWV), which is undergoing a worldwide epidemic, causes the deaths of individual honey bees and collapse of whole colonies. Three-dimensional structures of DWV determined at different conditions shows that the virus surface is decorated with protruding globular extensions of capsid proteins. The protruding domains contain a putative catalytic site that is probably required for the entry of the virus into the host cell. In addition, parts of the DWV RNA genome interact with the inside of the virus capsid. Identifying the RNA binding and catalytic sites within the DWV virion offers prospects for the development of antiviral treatments.

    Skubnik, K.; Novacek, J.; Füzik, T.; Pridal, A.; Paxton, R. J. & Plevka, P., Structure of deformed wing virus, a major honey bee pathogen, PNAS, 114, 3210-3215 (2017) DOI: 10.1073/pnas.1615695114

  • EMBO Rep. 2017

    EMBO Rep. 2017

    NMR structure shows how the CTD-interacting domain of Rtt103p recognizes threonine-4 phosphorylated CTD of RNA polymerase II (RNAPII).

    Richard Štefl Research Group


    Phosphorylation patterns of the C‐terminal domain (CTD) of largest subunit of RNApolymerase II (called the CTD code) orchestrate the recruitment of RNA processing and transcription factors. Recent studies showed that not only serines and tyrosines but also threonines of the CTD can be phosphorylated with a number of functional consequences, including the interaction with yeast transcription termination factor, Rtt103p. The solution structure of the Rtt103p CTD‐interacting domain (CID) bound to Thr4 phosphorylated CTD has been obtained by NMR. The structure reveals a direct recognition of the phospho‐Thr4 mark by Rtt103p CID and shows extensive interactions involving residues from three repeats of the CTD heptad. The structural data suggests that the recruitment of a CID‐containing CTD‐binding factor may be coded by more than one letter of the CTD code.

    Jasnovidova, O.; Krejcikova, M.; Kubicek, K. & Stefl, R. Structural insight into recognition of phosphorylated threonine-4 of RNA polymerase II C-terminal domain by Rtt103p. Embo Reports 18, 906-913, doi:10.15252/embr.201643723 (2017).

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