Journal of Structural Biology

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  • Biogenic nanospheres of amorphous carbonated Ca–Mg phosphate within the periostracum of the green mussel Perna viridis
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): Jun Xu , Gangsheng Zhang

    Recently there is increasing evidence that the shell biomineralization proceeds via an amorphous precursor route. Therefore, the search for and investigation of amorphous biominerals in bivalve shells are of great importance and interest. Here, using a scanning electron microscope (SEM), transmission electron microscope (TEM), X-ray diffractometer (XRD), and Fourier transform infrared spectrometer (FTIR), we investigate the microstructure and mineralogy of the periostracum in Perna viridis. We find that: (1) the periostracum has three layers, of which the inner and outer layer are of proteins, while the middle layer is mineralized with nanospheres of amorphous biominerals; (2) the nanospheres are of amorphous carbonated Ca–Mg phosphate (ACCP), where the CO3 2 −/PO4 3 − weight ratio is estimated to be ∼0.3, and the Ca/P and Ca/Mg atomic ratio is ∼1.4 and 1.6, respectively; (3) the nanospheres, with a diameter of 43–106nm, are found to assemble into spherules with a diameter of 160–500nm, which are further organized into parallel microlayers separated by the proteins; and (4) the nanospheres are assumed to function as the pH stabilizer to facilitate the shell’s initial mineralization. Finally, we expect that these findings will advance our understanding of the shell’s biomineralization process.





    Categories: Journal Articles
  • Amino acid sequence homologies in the hard keratins of birds and reptiles, and their implications for molecular structure and physical properties
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): R.D. Bruce Fraser , David A.D. Parry

    Avian and reptilian epidermal appendages such as feathers, claws and scales exhibit a filament–matrix texture. Previous studies have established that both components reside within the same single-chain molecule. In the present study the homology in a wide range of aligned sequences is used to gain insights into the structure and function of the molecular segments associated with the filament and with the matrix. The notion that all molecules contain a β-rich 34-residue segment associated with the framework of the filament is reinforced by the present study. In addition, the residues involved in the polymerization of the molecules to form filaments are identified. In the Archosaurs (birds, crocodiles and turtles), and the Squamates (snakes and lizards) segments rich in glycine and tyrosine can be identified in the C-terminal domain. In Rhynocephalians (tuataras) and Squamates a similar segment is inserted at a specific point in the N-terminal domain. In some Archosaurian appendages (both avian and reptilian) segments rich in charged residues and cysteine are found in the N-terminal domain. The likely effect of these segments will be to soften the tissue without compromising its insolubility. The structure and role of the various molecular segments identified in this study and the way in which they might manifest themselves in terms of the physical properties of the particular epidermal appendage in which they appear are also discussed.





    Categories: Journal Articles
  • Thalidomide mimics uridine binding to an aromatic cage in cereblon
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): Marcus D. Hartmann , Iuliia Boichenko , Murray Coles , Fabio Zanini , Andrei N. Lupas , Birte Hernandez Alvarez

    Thalidomide and its derivatives lenalidomide and pomalidomide are important anticancer agents but can cause severe birth defects via an interaction with the protein cereblon. The ligand-binding domain of cereblon is found, with a high degree of conservation, in both bacteria and eukaryotes. Using a bacterial model system, we reveal the structural determinants of cereblon substrate recognition, based on a series of high-resolution crystal structures. For the first time, we identify a cellular ligand that is universally present: we show that thalidomide and its derivatives mimic and compete for the binding of uridine, and validate these findings in vivo. The nature of the binding pocket, an aromatic cage of three tryptophan residues, further suggests a role in the recognition of cationic ligands. Our results allow for general evaluation of pharmaceuticals for potential cereblon-dependent teratogenicity.





    Categories: Journal Articles
  • Crystal structure of Legionella pneumophila dephospho-CoA kinase reveals a non-canonical conformation of P-loop
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): Xiaojian Gong , Xiaofang Chen , Dongmin Yu , Nannan Zhang , Zhongliang Zhu , Liwen Niu , Yuxin Mao , Honghua Ge

    Dephospho-CoA kinase (DPCK; EC 2.7.1.24) catalyzes the final step in the coenzyme A biosynthetic pathway. DPCK transfers a phosphate group from ATP to the 3-hydroxyl group of the ribose of dephosphocoenzyme A (dCoA) to yield CoA and ADP. Upon the binding of ligands, large conformational changes is induced in DPCKs, as well as in many other kinases, to shield the bound ATP in their catalytic site from the futile hydrolysis by bulk water molecules. To investigate the molecular mechanisms underlying the phosphoryl transfer during DPCK catalytic cycle, we determined the crystal structures of the Legionella pneumophila DPCK (LpDPCK) both in its apo-form and in complex with ATP. The structures reveal that LpDPCK comprises of three domains, the classical core domain, the CoA domain, and the LID domain, which are packed together to create a central cavity for substrate-binding and enzymatic catalysis. The binding of ATP induces large conformational changes, including a hinge-bending motion of the CoA binding domain and the “helix to loop” conformational change of the P-loop. Finally, modeling of a dCoA molecule to the enzyme provides insights into the catalytic mechanism of DPCK.





    Categories: Journal Articles
  • Dark-field transmission electron microscopy of cortical bone reveals details of extrafibrillar crystals
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): Henry P. Schwarcz , Elizabeth A. McNally , Gianluigi A. Botton

    In a previous study we showed that most of the mineral in bone is present in the form of “mineral structures”, 5–6nm-thick, elongated plates which surround and are oriented parallel to collagen fibrils. Using dark-field transmission electron microscopy, we viewed mineral structures in ion-milled sections of cortical human bone cut parallel to the collagen fibrils. Within the mineral structures we observe single crystals of apatite averaging 5.8±2.7nm in width and 28±19nm in length, their long axes oriented parallel to the fibril axis. Some appear to be composite, co-aligned crystals as thin as 2nm. From their similarity to TEM images of crystals liberated from deproteinated bone we infer that we are viewing sections through platy crystals of apatite that are assembled together to form the mineral structures.





    Categories: Journal Articles
  • Crystal structure of the VapBC-15 complex from Mycobacterium tuberculosis reveals a two-metal ion dependent PIN-domain ribonuclease and a variable mode of toxin–antitoxin assembly
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): Uddipan Das , Vivian Pogenberg , Udaya Kumar Tiruttani Subhramanyam , Matthias Wilmanns , Samudrala Gourinath , Alagiri Srinivasan

    Although PIN (PilT N-terminal)-domain proteins are known to have ribonuclease activity, their specific mechanism of action remains unknown. VapCs form a family of ribonucleases that possess a PIN-domain assembly and are known as toxins. The activities of VapCs are impaired by VapB antitoxins. Here we present the crystal structure of the VapBC-15 toxin–antitoxin complex from Mycobacterium tuberculosis determined to 2.1Å resolution. The VapB-15 and VapC-15 components assemble into one heterotetramer (VapB2C2) and two heterotrimers (VapBC2) in each asymmetric unit of the crystal. The active site of VapC-15 toxin consists of a cluster of acidic amino acid residues and two divalent metal ions, forming a well organised ribonuclease active site. The distribution of the catalytic-site residues of the VapC-15 toxin is similar to that of T4 RNase H and of Methanococcus jannaschii FEN-1, providing strong evidence that these three proteins share a similar mechanism of activity. The presence of both VapB2C2 and VapBC2 emphasizes the fact that the same antitoxin can bind the toxin in 1:1 and 1:2 ratios. The crystal structure determination of the VapBC-15 complex reveals for the first time a PIN-domain ribonuclease protein that shows two metal ions at the active site and a variable mode of toxin–antitoxin assembly. The structure further shows that VapB-15 antitoxin binds to the same groove meant for the binding of putative substrate (RNA), resulting in the inhibition of VapC-15’s toxicity.





    Categories: Journal Articles
  • Separation of replication and transcription domains in nucleoli
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): E. Smirnov , J. Borkovec , L. Kováčik , S. Svidenská , A. Schröfel , M. Skalníková , Z. Švindrych , P. Křížek , M. Ovesný , G.M. Hagen , P. Juda , K. Michalová , M.V. Cardoso , D. Cmarko , I. Raška

    In mammalian cells, active ribosomal genes produce the 18S, 5.8S and 28S RNAs of ribosomal particles. Transcription levels of these genes are very high throughout interphase, and the cell needs a special strategy to avoid collision of the DNA polymerase and RNA polymerase machineries. To investigate this problem, we measured the correlation of various replication and transcription signals in the nucleoli of HeLa, HT-1080 and NIH 3T3 cells using a specially devised software for analysis of confocal images. Additionally, to follow the relationship between nucleolar replication and transcription in living cells, we produced a stable cell line expressing GFP-RPA43 (subunit of RNA polymerase I, pol I) and RFP-PCNA (the sliding clamp protein) based on human fibrosarcoma HT-1080 cells. We found that replication and transcription signals are more efficiently separated in nucleoli than in the nucleoplasm. In the course of S phase, separation of PCNA and pol I signals gradually increased. During the same period, separation of pol I and incorporated Cy5-dUTP signals decreased. Analysis of single molecule localization microscopy (SMLM) images indicated that transcriptionally active FC/DFC units (i.e. fibrillar centers with adjacent dense fibrillar components) did not incorporate DNA nucleotides. Taken together, our data show that replication of the ribosomal genes is spatially separated from their transcription, and FC/DFC units may provide a structural basis for that separation.





    Categories: Journal Articles
  • Particle migration analysis in iterative classification of cryo-EM single-particle data
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): Bo Chen , Bingxin Shen , Joachim Frank

    Recently developed classification methods have enabled resolving multiple biological structures from cryo-EM data collected on heterogeneous biological samples. However, there remains the problem of how to base the decisions in the classification on the statistics of the cryo-EM data, to reduce the subjectivity in the process. Here, we propose a quantitative analysis to determine the iteration of convergence and the number of distinguishable classes, based on the statistics of the single particles in an iterative classification scheme. We start the classification with more number of classes than anticipated based on prior knowledge, and then combine the classes that yield similar reconstructions. The classes yielding similar reconstructions can be identified from the migrating particles (jumpers) during consecutive iterations after the iteration of convergence. We therefore termed the method “jumper analysis”, and applied it to the output of RELION 3D classification of a benchmark experimental dataset. This work is a step forward toward fully automated single-particle reconstruction and classification of cryo-EM data.





    Categories: Journal Articles
  • Crystal structure of the essential Mycobacterium tuberculosis phosphopantetheinyl transferase PptT, solved as a fusion protein with maltose binding protein
    [Feb 2015]

    Publication date: December 2014
    Source:Journal of Structural Biology, Volume 188, Issue 3

    Author(s): James Jung , Ghader Bashiri , Jodie M. Johnston , Alistair S. Brown , David F. Ackerley , Edward N. Baker

    Phosphopantetheinyl transferases (PPTases) are key enzymes in the assembly-line production of complex molecules such as fatty acids, polyketides and polypeptides, where they activate acyl or peptidyl carrier proteins, transferring a 4′-phosphopantetheinyl moiety from coenzyme A (CoA) to a reactive serine residue on the carrier protein. The human pathogen Mycobacterium tuberculosis encodes two PPTases, both essential and therefore attractive drug targets. We report the structure of the type-II PPTase PptT, obtained from crystals of a fusion protein with maltose binding protein. The structure, at 1.75Å resolution (R =0.156, R free =0.191), reveals an α/β fold broadly similar to other type-II PPTases, but with differences in peripheral structural elements. A bound CoA is clearly defined with its pantetheinyl arm tucked into a hydrophobic pocket. Interactions involving the CoA diphosphate, bound Mg2+ and three active site acidic side chains suggest a plausible pathway for proton transfer during catalysis.





    Categories: Journal Articles
  • Cover 2 - Editorial Board
    [Feb 2015]

    Publication date: November 2014
    Source:Journal of Structural Biology, Volume 188, Issue 2









    Categories: Journal Articles
  • DNA buckling in bacteriophage cavities as a mechanism to aid virus assembly
    [Jan 2015]

    Publication date: Available online 19 January 2015
    Source:Journal of Structural Biology

    Author(s): Andrew D. Hirsh , N.C. Perkins

    While relatively simple biologically, bacteriophages are sophisticated biochemical machines that execute a precise sequence of events during virus assembly, DNA packaging, and ejection. These stages of the viral life cycle require intricate coordination of viral components whose structures are being revealed by single molecule experiments and high resolution (cryo-electron microscopy) reconstructions. For example, during packaging, bacteriophages employ some of the strongest known molecular motors to package DNA against increasing pressure within the viral capsid shell. Located upstream of the motor is an elaborate portal system through which DNA is threaded. A high resolution reconstruction of the portal system for bacteriophage ϕ 29 reveals that DNA buckles inside a small cavity under large compressive forces. In this study, we demonstrate that DNA can also buckle in other bacteriophages including T7 and P22. Using a computational rod model for DNA, we demonstrate that a DNA buckle can initiate and grow within the small confines of a cavity under biologically-attainable force levels. The forces of DNA-cavity contact and DNA-DNA electrostatic repulsion ultimately limit cavity filling. Despite conforming to very different cavity geometries, the buckled DNA within T7 and P22 exhibits near equal volumetric energy density (∼ 1 kT/ nm 3 ) and energetic cost of packaging (∼ 22 kT). We hypothesize that a DNA buckle creates large forces on the cavity interior to signal the conformational changes to end packaging. In addition, a DNA buckle may help retain the genome prior to tail assembly through significantly increased contact area with the portal.





    Categories: Journal Articles
  • Paper of the Year Award
    [Jan 2015]

    Publication date: Available online 17 January 2015
    Source:Journal of Structural Biology









    Categories: Journal Articles
  • Paper of the Year Award
    [Jan 2015]

    Publication date: Available online 17 January 2015
    Source:Journal of Structural Biology









    Categories: Journal Articles
  • Corrigendum to “A clarification of the terms used in comparing semi-automated particle selection algorithms in cryo-EM” [J. Struct. Biol. 175 (2011) 348–352]
    [Jan 2015]

    Publication date: Available online 16 January 2015
    Source:Journal of Structural Biology

    Author(s): Robert Langlois , Joachim Frank







    Categories: Journal Articles
  • Evidence of compositional and ultrastructural shifts during the development of calcareous tubes in the biofouling tubeworm, Hydroides elegans
    [Jan 2015]

    Publication date: Available online 16 January 2015
    Source:Journal of Structural Biology

    Author(s): Vera Bin San Chan , Olev Vinn , Chaoyi Li , Xingwen Lu , Anatoliy B. Kudryavtsev , J. William Schopf , Kaimin Shih , Tong Zhang , Vengatesen Thiyagarajan

    The serpulid tubeworm, Hydroides elegans, is an ecologically and economically important species whose biology has been fairly well studied, especially in the context of larval development and settlement on man-made objects (biofouling). Nevertheless, ontogenetic changes associated with calcareous tube composition and structures have not yet been studied. Here, the ultrastructure and composition of the calcareous tubes built by H. elegans was examined in the three early calcifying juvenile stages and in the adult using XRD, FTIR, ICP-OES, SEM and Raman spectroscopy. Ontogenetic shifts in carbonate mineralogy were observed, for example, juvenile tubes contained more amorphous calcium carbonate and were predominantly aragonitic whereas adult tubes were bimineralic with considerably more calcite. The mineral composition gradually shifted during the tube development as shown by a decrease in Sr/Ca and an increase of Mg/Ca ratios with the tubeworm’s age. The inner tube layer contained calcite, whereas the outer layer contained aragonite. Similarly, the tube complexity in terms of ultrastructure was associated with development. The sequential appearance of unoriented ultrastructures followed by oriented ultrastructures may reflect the evolutionary history of serpulid tube biominerals. As aragonitic structures are more susceptible to dissolution under ocean acidification (OA) conditions but are more difficult to be removed by anti-fouling treatments, the early developmental stages of the tubeworms may be vulnerable to OA but act as the important target for biofouling control.





    Categories: Journal Articles
  • Structural and functional analysis of two universal stress proteins YdaA and YnaF from Salmonella typhimurium: Possible roles in microbial stress tolerance
    [Jan 2015]

    Publication date: Available online 16 January 2015
    Source:Journal of Structural Biology

    Author(s): M. Bangera , R. Panigrahi , S.R. Sagurthi , H.S. Savithri , M.R.N. Murthy

    In many organisms “universal stress proteins” (USPs) are induced in response to a variety of environmental stresses. Here we report the structures of two USPs, YnaF and YdaA from Salmonella typhimurium determined at 1.8Å and 2.4Å resolutions, respectively. YnaF consists of a single USP domain and forms a tetrameric organization stabilized by interactions mediated through chloride ions. YdaA is a larger protein consisting of two tandem USP domains. Two protomers of YdaA associate to form a structure similar to the YnaF tetramer. YdaA showed ATPase activity and an ATP binding motif G-2X-G-9X-G(S/T/N) was found in its C-terminal domain. The residues corresponding to this motif were not conserved in YnaF although YnaF could bind ATP. However, unlike YdaA, YnaF did not hydrolyse ATP in vitro. Disruption of interactions mediated through chloride ions by selected mutations converted YnaF into an ATPase. Residues that might be important for ATP hydrolysis could be identified by comparing the active sites of native and mutant structures. Only the C-terminal domain of YdaA appears to be involved in ATP hydrolysis. The structurally similar N-terminal domain was found to bind a zinc ion near the segment equivalent to the phosphate binding loop of the C-terminal domain. Mass spectrometric analysis showed that YdaA might bind a ligand of approximate molecular weight 800Daltons. Structural comparisons suggest that the ligand, probably related to an intermediate in lipid A biosynthesis, might bind at a site close to the zinc ion. Therefore, the N-terminal domain of YdaA binds zinc and might play a role in lipid metabolism. Thus, USPs appear to perform several distinct functions such as ATP hydrolysis, altering membrane properties and chloride sensing.





    Categories: Journal Articles
  • Crystal structures for short-chain pentraxin from zebrafish demonstrate a cyclic trimer with new recognition and effector faces
    [Jan 2015]

    Publication date: Available online 13 January 2015
    Source:Journal of Structural Biology

    Author(s): Rong Chen , Jianxun Qi , Hongyu Yuan , Yanan Wu , Wei Hu , Chun Xia

    Short-chain pentraxins (PTXs), including CRP and SAP, are innate pattern recognition receptors that play vital roles in the recognition and elimination of various pathogenic bacteria by triggering the classical complement pathway through C1q. Similar to antibodies, pentraxins can also activate opsonisation and phagocytosis by interacting with Fc receptors (FcRs). Various structural studies on human PTXs have been performed, but there are no reports about the crystal structure of bony fish pentraxins. Here, the crystal structures of zebrafish PTX (Dare-PTX-Ca and Dare-PTX) are presented. Both Dare-PTX-Ca and Dare-PTX are cyclic trimers, which are new forms of crystallised pentraxins. The structures reveal that the ligand-binding pocket (LBP) in the recognition face of Dare-PTX is deep and narrow. Homology modelling shows that LBPs from different Dare-PTX loci differ in shape, reflecting their specific recognition abilities. Furthermore, in comparison with the structure of hCPR, a new C1q binding mode was identified in Dare-PTX. In addition, the FcR-binding sites of hSAP are partially conserved in Dare-PTX. These results will shed light on the understanding of a primitive PTX in bony fish, which evolved approximately 450 million years ago.





    Categories: Journal Articles
  • Structure of inorganic pyrophosphatase from Staphylococcus aureus reveals conformational flexibility of the active site
    [Jan 2015]

    Publication date: Available online 7 January 2015
    Source:Journal of Structural Biology

    Author(s): Chathurada S. Gajadeera , Xinyi Zhang , Yinan Wei , Oleg V. Tsodikov

    Cytoplasmic inorganic pyrophosphatase (PPiase) is an enzyme essential for survival of organisms, from bacteria to human. PPiases are divided into two structurally distinct families: family I PPiases are Mg2+-dependent and present in most archaea, eukaryotes and prokaryotes, whereas the relatively less understood family II PPiases are Mn2+-dependent and present only in some archaea, bacteria and primitive eukaryotes. Staphylococcus aureus (SA), a dangerous pathogen and a frequent cause of hospital infections, contains a family II PPiase (PpaC), which is an attractive potential target for development of novel antibacterial agents. We determined a crystal structure of SA PpaC in complex with catalytic Mn2+ at 2.1Å resolution. The active site contains two catalytic Mn2+ binding sites, each half-occupied, reconciling the previously observed 1:1 Mn2+:enzyme stoichiometry with the presence of two divalent metal ion sites in the apo-enzyme. Unexpectedly, despite the absence of the substrate or products in the active site, the two domains of SA PpaC form a closed active site, a conformation observed in structures of other family II PPiases only in complex with substrate or product mimics. A region spanning residues 295–298, which contains a conserved substrate binding RKK motif, is flipped out of the active site, an unprecedented conformation for a PPiase. Because the mutant of Arg295 to an alanine is devoid of activity, this loop likely undergoes an induced-fit conformational change upon substrate binding and product dissociation. This closed conformation of SA PPiase may serve as an attractive target for rational design of inhibitors of this enzyme.





    Categories: Journal Articles
  • Cover 2 - Editorial Board
    [Jan 2015]

    Publication date: January 2015
    Source:Journal of Structural Biology, Volume 189, Issue 1









    Categories: Journal Articles
  • Table of Contents / barcode
    [Jan 2015]

    Publication date: January 2015
    Source:Journal of Structural Biology, Volume 189, Issue 1









    Categories: Journal Articles