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https://www.readbyqxmd.com/read/28732205/unique-roles-of-the-non-identical-mcm-subunits-in-dna-replication-licensing
#1
REVIEW
Yuanliang Zhai, Ningning Li, Hanlun Jiang, Xuhui Huang, Ning Gao, Bik Kwoon Tye
A family of six homologous subunits, Mcm2, -3, -4, -5, -6, and -7, each with its own unique features, forms the catalytic core of the eukaryotic replicative helicase. The necessity of six similar but non-identical subunits has been a mystery since its initial discovery. Recent cryo-EM structures of the Mcm2-7 (MCM) double hexamer, its precursors, and the origin recognition complex (ORC)-Cdc6-Cdt1-Mcm2-7 (OCCM) intermediate showed that each of these subunits plays a distinct role in orchestrating the assembly of the pre-replication complex (pre-RC) by ORC-Cdc6 and Cdt1...
July 20, 2017: Molecular Cell
https://www.readbyqxmd.com/read/28731030/cryo-em-structure-and-biochemical-analysis-reveal-the-basis-of-the-functional-difference-between-human-pi3kc3-c1-and-c2
#2
Meisheng Ma, Jun-Jie Liu, Yan Li, Yuwei Huang, Na Ta, Yang Chen, Hua Fu, Ming-Da Ye, Yuehe Ding, Weijiao Huang, Jia Wang, Meng-Qiu Dong, Li Yu, Hong-Wei Wang
Phosphatidylinositol 3-phosphate (PI3P) plays essential roles in vesicular trafficking, organelle biogenesis and autophagy. Two class III phosphatidylinositol 3-kinase (PI3KC3) complexes have been identified in mammals, the ATG14L complex (PI3KC3-C1) and the UVRAG complex (PI3KC3-C2). PI3KC3-C1 is crucial for autophagosome biogenesis, and PI3KC3-C2 is involved in various membrane trafficking events. Here we report the cryo-EM structures of human PI3KC3-C1 and PI3KC3-C2 at sub-nanometer resolution. The two structures share a common L-shaped overall architecture with distinct features...
July 21, 2017: Cell Research
https://www.readbyqxmd.com/read/28716758/oral-delivery-of-vancomycin-by-tetraether-lipid-liposomes
#3
Philipp Uhl, Silvia Pantze, Philip Storck, Johannes Parmentier, Dominik Witzigmann, Götz Hofhaus, Jörg Huwyler, Walter Mier, Gert Fricker
Despite the outstanding progress in modern medicine, the oral delivery of peptide drugs is limited until today due to their instability in the gastrointestinal tract and low mucosa penetration. To overcome these hurdles, liposomes containing the specific tetraether lipid GCTE (glycerylcaldityltetraether lipid) were examined. For this purpose, the glycopeptide antibiotic vancomycin was used as model substance and liposomes were prepared by DAC (dual assymetric centrifugation). These liposomes showed a size and polydispersity index comparable to standard liposomes...
July 14, 2017: European Journal of Pharmaceutical Sciences
https://www.readbyqxmd.com/read/28714467/cryo-em-structures-of-the-atp-bound-vps4-e233q-hexamer-and-its-complex-with-vta1-at-near-atomic-resolution
#4
Shan Sun, Lin Li, Fan Yang, Xiaojing Wang, Fenghui Fan, Mengyi Yang, Chunlai Chen, Xueming Li, Hong-Wei Wang, Sen-Fang Sui
The cellular ESCRT-III (endosomal sorting complex required for transport-III) and Vps4 (vacuolar protein sorting 4) comprise a common machinery that mediates a variety of membrane remodelling events. Vps4 is essential for the machinery function by using the energy from ATP hydrolysis to disassemble the ESCRT-III polymer into individual proteins. Here, we report the structures of the ATP-bound Vps4(E233Q) hexamer and its complex with the cofactor Vta1 (vps twenty associated 1) at resolutions of 3.9 and 4.2 Å, respectively, determined by electron cryo-microscopy...
July 17, 2017: Nature Communications
https://www.readbyqxmd.com/read/28711156/towards-a-structural-view-of-drug-binding-to-herg-k-channels
#5
REVIEW
Jamie I Vandenberg, Eduardo Perozo, Toby W Allen
The human ether-a-go-go-related gene (hERG) K(+) channel is of great medical and pharmaceutical relevance. Inherited mutations in hERG result in congenital long-QT syndrome which is associated with a markedly increased risk of cardiac arrhythmia and sudden death. hERG K(+) channels are also remarkably susceptible to block by a wide range of drugs, which in turn can cause drug-induced long-QT syndrome and an increased risk of sudden death. The recent determination of the near-atomic resolution structure of the hERG K(+) channel, using single-particle cryo-electron microscopy (cryo-EM), provides tremendous insights into how these channels work...
July 12, 2017: Trends in Pharmacological Sciences
https://www.readbyqxmd.com/read/28710447/cryo-em-reconstruction-of-the-cafeteria-roenbergensis-virus-capsid-suggests-novel-assembly-pathway-for-giant-viruses
#6
Chuan Xiao, Matthias G Fischer, Duer M Bolotaulo, Nancy Ulloa-Rondeau, Gustavo A Avila, Curtis A Suttle
Whereas the protein composition and overall shape of several giant virus capsids have been described, the mechanism by which these large capsids assemble remains enigmatic. Here, we present a reconstruction of the capsid of Cafeteria roenbergensis virus (CroV), one of the largest viruses analyzed by cryo-electron microscopy (cryo-EM) to date. The CroV capsid has a diameter of 3,000 Å and a Triangulation number of 499. Unlike related mimiviruses, the CroV capsid is not decorated with glycosylated surface fibers, but features 30 Å-long surface protrusions that are formed by loops of the major capsid protein...
July 14, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28710336/subunit-conformational-variation-within-individual-groel-oligomers-resolved-by-cryo-em
#7
Soung-Hun Roh, Corey F Hryc, Hyun-Hwan Jeong, Xue Fei, Joanita Jakana, George H Lorimer, Wah Chiu
Single-particle electron cryo-microscopy (cryo-EM) is an emerging tool for resolving structures of conformationally heterogeneous particles; however, each structure is derived from an average of many particles with presumed identical conformations. We used a 3.5-Å cryo-EM reconstruction with imposed D7 symmetry to further analyze structural heterogeneity among chemically identical subunits in each GroEL oligomer. Focused classification of the 14 subunits in each oligomer revealed three dominant classes of subunit conformations...
July 14, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28692650/expression-purification-and-contaminant-detection-for-structural-studies-of-ralstonia-metallidurance-clc-protein-rm1
#8
Priyanka D Abeyrathne, Nikolaus Grigorieff
Single-particle electron cryo-microscopy (cryo-EM) has become a popular method for high-resolution study of the structural and functional properties of proteins. However, sufficient expression and purification of membrane proteins holds many challenges. We describe methods to overcome these obstacles using ClC-rm1, a prokaryotic chloride channel (ClC) family protein from Ralstonia metallidurans, overexpressed in Escherichia coli (E. coli) BL21(DE3) strain. Mass spectrometry and electron microscopy analyses of purified samples revealed multiple contaminants that can obfuscate results of subsequent high-resolution structural analysis...
2017: PloS One
https://www.readbyqxmd.com/read/28683309/the-complete-structure-of-the-mycobacterium-smegmatis-70s-ribosome
#9
Jendrik Hentschel, Chloe Burnside, Ingrid Mignot, Marc Leibundgut, Daniel Boehringer, Nenad Ban
The ribosome carries out the synthesis of proteins in every living cell. It consequently represents a frontline target in anti-microbial therapy. Tuberculosis ranks among the leading causes of death worldwide, due in large part to the combination of difficult-to-treat latency and antibiotic resistance. Here, we present the 3.3-Å cryo-EM structure of the 70S ribosome of Mycobacterium smegmatis, a close relative to the human pathogen Mycobacterium tuberculosis. The structure reveals two additional ribosomal proteins and localizes them to the vicinity of drug-target sites in both the catalytic center and the decoding site of the ribosome...
July 5, 2017: Cell Reports
https://www.readbyqxmd.com/read/28682307/cryo-em-structure-of-the-protein-conducting-erad-channel-hrd1-in-complex-with-hrd3
#10
Stefan Schoebel, Wei Mi, Alexander Stein, Sergey Ovchinnikov, Ryan Pavlovicz, Frank DiMaio, David Baker, Melissa G Chambers, Huayou Su, Dongsheng Li, Tom A Rapoport, Maofu Liao
Misfolded endoplasmic reticulum (ER) proteins are retro-translocated through the membrane into the cytosol, where they are poly-ubiquitinated, extracted from the ER membrane, and degraded by the proteasome(1-4), a pathway termed ER-associated protein degradation (ERAD). Proteins with misfolded domains in the ER lumen or membrane are discarded through the ERAD-L and -M pathways, respectively. In S. cerevisiae, both pathways require the ubiquitin ligase Hrd1, a multi-spanning membrane protein with a cytosolic RING finger domain(5,6)...
July 6, 2017: Nature
https://www.readbyqxmd.com/read/28678775/cryo-em-structures-of-tau-filaments-from-alzheimer-s-disease
#11
Anthony W P Fitzpatrick, Benjamin Falcon, Shaoda He, Alexey G Murzin, Garib Murshudov, Holly J Garringer, R Anthony Crowther, Bernardino Ghetti, Michel Goedert, Sjors H W Scheres
Alzheimer's disease is the most common neurodegenerative disease, and there are no mechanism-based therapies. The disease is defined by the presence of abundant neurofibrillary lesions and neuritic plaques in the cerebral cortex. Neurofibrillary lesions comprise paired helical and straight tau filaments, whereas tau filaments with different morphologies characterize other neurodegenerative diseases. No high-resolution structures of tau filaments are available. Here we present cryo-electron microscopy (cryo-EM) maps at 3...
July 13, 2017: Nature
https://www.readbyqxmd.com/read/28675816/expanding-the-boundaries-of-cryo-em-with-phase-plates
#12
REVIEW
Radostin Danev, Wolfgang Baumeister
Phase plates have long been considered as a means for improving the performance of cryo-electron microscopy (cryo-EM). But practical limitations, such as a short lifespan or cumbersome usage have prevented their widespread adoption. The recently developed Volta phase plate overcomes most of the practical issues and it is now commercially available. Results from both, electron cryo-tomography (cryo-ET) and single particle analysis (SPA), have demonstrated the benefits of using a phase plate. In CET phase plates have helped to visualize cellular ultrastructure in unprecedented detail...
July 1, 2017: Current Opinion in Structural Biology
https://www.readbyqxmd.com/read/28671674/addressing-preferred-specimen-orientation-in-single-particle-cryo-em-through-tilting
#13
Yong Zi Tan, Philip R Baldwin, Joseph H Davis, James R Williamson, Clinton S Potter, Bridget Carragher, Dmitry Lyumkis
We present a strategy for tackling preferred specimen orientation in single-particle cryogenic electron microscopy by employing tilts during data collection. We also describe a tool to quantify the resulting directional resolution using 3D Fourier shell correlation volumes. We applied these methods to determine the structures at near-atomic resolution of the influenza hemagglutinin trimer, which adopts a highly preferred specimen orientation, and of ribosomal biogenesis intermediates, which adopt moderately preferred orientations...
July 3, 2017: Nature Methods
https://www.readbyqxmd.com/read/28671286/structural-characterization-of-the-nap-the-major-adhesion-complex-of-the-human-pathogen-mycoplasma-genitalium
#14
Margot P Scheffer, Luis Gonzalez-Gonzalez, Anja Seybert, Mercè Ratera, Michael Kunz, José M Valpuesta, Ignacio Fita, Enrique Querol, Jaume Piñol, Jaime Martín-Benito, Achilleas S Frangakis
Mycoplasma genitalium, the causative agent of non-gonococcal urethritis and pelvic inflammatory disease in humans, is a small eubacterium that lacks a peptidoglycan cell wall. On the surface of its plasma membrane is the major surface adhesion complex, known as NAP, that is essential for adhesion and gliding motility of the organism. Here, we have performed cryo-electron tomography of intact cells and detergent permeabilized M. genitalium cell aggregates, providing sub-tomogram averages of free and cell-attached NAPs respectively, revealing a tetrameric complex with two-fold rotational (C2) symmetry...
July 3, 2017: Molecular Microbiology
https://www.readbyqxmd.com/read/28667626/structural-analysis-of-protein-complexes-by-cryo-electron-microscopy
#15
Tiago R D Costa, Athanasios Ignatiou, Elena V Orlova
Structural studies of biocomplexes using single-particle cryo-electron microscopy (cryo-EM) is now a well-established technique in structural biology and has become competitive with X-ray crystallography. The latest advances in EM enable us to determine structures of protein complexes at 3-5 Å resolution for an extremely broad range of sizes from ~200 kDa up to hundreds of megadaltons (Bartesaghi et al., Science 348(6239):1147-1151, 2051; Bai et al., Nature 525(7568):212-217, 2015; Vinothkumar et al., Nature 515(7525):80-84, 2014; Grigorieff and Harrison, Curr Opin Struct Biol 21(2):265-273, 2011)...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28666122/structure-basis-for-directional-r-loop-formation-and-substrate-handover-mechanisms-in-type-i-crispr-cas-system
#16
Yibei Xiao, Min Luo, Robert P Hayes, Jonathan Kim, Sherwin Ng, Fang Ding, Maofu Liao, Ailong Ke
Type I CRISPR systems feature a sequential dsDNA target searching and degradation process, by crRNA-displaying Cascade and nuclease-helicase fusion enzyme Cas3, respectively. Here we present two cryo-EM snapshots of the Thermobifida fusca type I-E Cascade: (1) unwinding 11 bp of dsDNA at the seed-sequence region to scout for sequence complementarity, and (2) further unwinding of the entire protospacer to form a full R-loop. These structures provide the much-needed temporal and spatial resolution to resolve key mechanistic steps leading to Cas3 recruitment...
June 29, 2017: Cell
https://www.readbyqxmd.com/read/28665412/cryo-em-structure-of-haemoglobin-at-3-2-%C3%A3-determined-with-the-volta-phase-plate
#17
Maryam Khoshouei, Mazdak Radjainia, Wolfgang Baumeister, Radostin Danev
With the advent of direct electron detectors, the perspectives of cryo-electron microscopy (cryo-EM) have changed in a profound way. These cameras are superior to previous detectors in coping with the intrinsically low contrast and beam-induced motion of radiation-sensitive organic materials embedded in amorphous ice, and hence they have enabled the structure determination of many macromolecular assemblies to atomic or near-atomic resolution. Nevertheless, there are still limitations and one of them is the size of the target structure...
June 30, 2017: Nature Communications
https://www.readbyqxmd.com/read/28657312/molecular-dynamics-flexible-fitting-simulations-identify-new-models-of-the-closed-state-of-the-cystic-fibrosis-transmembrane-conductance-regulator-protein
#18
Luba Simhaev, Nael A McCarty, Robert C Ford, Hanoch Senderowitz
Cystic fibrosis (CF) is a lethal, genetic disease found in particular in humans of European origin which is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel. The search for CF therapies acting by modulating the impaired function of mutant CFTR will be greatly advanced by high resolution structures of CFTR in different states. To date, two medium resolution electron microscopy (EM) structures of CFTR are available (one of a distant zebrafish (Danio rerio) CFTR ortholog and one of human CFTR)...
July 18, 2017: Journal of Chemical Information and Modeling
https://www.readbyqxmd.com/read/28652389/structural-differences-between-yeast-and-mammalian-microtubules-revealed-by-cryo-em
#19
Stuart C Howes, Elisabeth A Geyer, Benjamin LaFrance, Rui Zhang, Elizabeth H Kellogg, Stefan Westermann, Luke M Rice, Eva Nogales
Microtubules are polymers of αβ-tubulin heterodimers essential for all eukaryotes. Despite sequence conservation, there are significant structural differences between microtubules assembled in vitro from mammalian or budding yeast tubulin. Yeast MTs were not observed to undergo compaction at the interdimer interface as seen for mammalian microtubules upon GTP hydrolysis. Lack of compaction might reflect slower GTP hydrolysis or a different degree of allosteric coupling in the lattice. The microtubule plus end-tracking protein Bim1 binds yeast microtubules both between αβ-tubulin heterodimers, as seen for other organisms, and within tubulin dimers, but binds mammalian tubulin only at interdimer contacts...
June 26, 2017: Journal of Cell Biology
https://www.readbyqxmd.com/read/28652322/cryo-em-structure-of-the-dna-pk-holoenzyme
#20
Humayun Sharif, Yang Li, Yuanchen Dong, Liyi Dong, Wei Li Wang, Youdong Mao, Hao Wu
DNA-dependent protein kinase (DNA-PK) is a large protein complex central to the nonhomologous end joining (NHEJ) DNA-repair pathway. It comprises the DNA-PK catalytic subunit (DNA-PKcs) and the heterodimer of DNA-binding proteins Ku70 and Ku80. Here, we report the cryo-electron microscopy (cryo-EM) structures of human DNA-PKcs at 4.4-Å resolution and the DNA-PK holoenzyme at 5.8-Å resolution. The DNA-PKcs structure contains three distinct segments: the N-terminal region with an arm and a bridge, the circular cradle, and the head that includes the kinase domain...
July 11, 2017: Proceedings of the National Academy of Sciences of the United States of America
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