scholarly journals New Structural Insights into the Genome and Minor Capsid Proteins of BK Polyomavirus using Cryo-Electron Microscopy

Structure ◽  
2016 ◽  
Vol 24 (4) ◽  
pp. 528-536 ◽  
Author(s):  
Daniel L. Hurdiss ◽  
Ethan L. Morgan ◽  
Rebecca F. Thompson ◽  
Emma L. Prescott ◽  
Margarita M. Panou ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Capsid Proteins ◽  
Cryo Electron Microscopy ◽  
Bk Polyomavirus ◽  
Structural Insights

scholarly journals The 4.4 Å structure of the giant Melbournevirus virion belonging to the Marseilleviridae family

2021 ◽  
Author(s):  
Raymond N Burton-Smith ◽  
Hemanth K N Reddy ◽  
Martin Svenda ◽  
Chantal Abergel ◽  
Kenta Okamoto ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Capsid Protein ◽  
Major Capsid Protein ◽  
Capsid Proteins ◽  
Atomic Model ◽  
Penton Base ◽  
Cryo Electron Microscopy ◽  
Internal Membrane ◽  
Structural Details ◽  
Giant Viruses

Members of Marseilleviridae, one family of icosahedral giant viruses classified in 2012 have been identified worldwide in all types of environments. The virion shows a characteristic internal membrane extrusion at the five-fold vertices of the capsid, but its structural details need to be elucidated. We now report the 4.4 Å cryo-electron microscopy structure of the Melbournevirus capsid. An atomic model of the major capsid protein (MCP) shows a unique cup structure on the trimer that accommodates additional proteins. A polyalanine model of the penton base protein shows internally extended N- and C-terminals, which indirectly connect to the internal membrane extrusion. The Marseilleviruses share the same orientational organisation of the MCPs as PBCV-1 and CroV, but do not appear to possess a protein akin to the ″tape measure″ of these viruses. Minor capsid proteins named PC-β, zipper, and scaffold are proposed to control the dimensions of the capsid during assembly.

scholarly journals Cryo-electron Microscopy Study of the Genome Release of the Dicistrovirus Israeli Acute Bee Paralysis Virus

2016 ◽  
Vol 91 (4) ◽  
Author(s):  
Edukondalu Mullapudi ◽  
Tibor Füzik ◽  
Antonín Přidal ◽  
Pavel Plevka
Keyword(s):  
Electron Microscopy ◽  
The United States ◽  
Capsid Proteins ◽  
Content Type ◽  
Cryo Electron Microscopy ◽  
The Family ◽  
Electron Microscopy Analysis ◽  
Colony Collapse ◽  
Acute Bee Paralysis Virus ◽  
Paralysis Virus

ABSTRACT Viruses of the family Dicistroviridae can cause substantial economic damage by infecting agriculturally important insects. Israeli acute bee paralysis virus (IAPV) causes honeybee colony collapse disorder in the United States. High-resolution molecular details of the genome delivery mechanism of dicistroviruses are unknown. Here we present a cryo-electron microscopy analysis of IAPV virions induced to release their genomes in vitro. We determined structures of full IAPV virions primed to release their genomes to a resolution of 3.3 Å and of empty capsids to a resolution of 3.9 Å. We show that IAPV does not form expanded A particles before genome release as in the case of related enteroviruses of the family Picornaviridae. The structural changes observed in the empty IAPV particles include detachment of the VP4 minor capsid proteins from the inner face of the capsid and partial loss of the structure of the N-terminal arms of the VP2 capsid proteins. Unlike the case for many picornaviruses, the empty particles of IAPV are not expanded relative to the native virions and do not contain pores in their capsids that might serve as channels for genome release. Therefore, rearrangement of a unique region of the capsid is probably required for IAPV genome release. IMPORTANCE Honeybee populations in Europe and North America are declining due to pressure from pathogens, including viruses. Israeli acute bee paralysis virus (IAPV), a member of the family Dicistroviridae, causes honeybee colony collapse disorder in the United States. The delivery of virus genomes into host cells is necessary for the initiation of infection. Here we present a structural cryo-electron microscopy analysis of IAPV particles induced to release their genomes. We show that genome release is not preceded by an expansion of IAPV virions as in the case of related picornaviruses that infect vertebrates. Furthermore, minor capsid proteins detach from the capsid upon genome release. The genome leaves behind empty particles that have compact protein shells.

scholarly journals Structural insights into thermostabilization of leucine dehydrogenase from its atomic structure by cryo-electron microscopy

2019 ◽  
Vol 205 (1) ◽  
pp. 11-21 ◽  
Author(s):  
Hiroki Yamaguchi ◽  
Akiko Kamegawa ◽  
Kunio Nakata ◽  
Tatsuki Kashiwagi ◽  
Toshimi Mizukoshi ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Atomic Structure ◽  
Leucine Dehydrogenase ◽  
Cryo Electron Microscopy ◽  
Structural Insights

scholarly journals Structural insights into stressosome assembly

IUCrJ ◽  
2019 ◽  
Vol 6 (5) ◽  
pp. 938-947 ◽  
Author(s):  
Eunju Kwon ◽  
Deepak Pathak ◽  
Han-ul Kim ◽  
Pawan Dahal ◽  
Sung Chul Ha ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Electron Microscopy ◽  
Environmental Stress ◽  
Assembly Model ◽  
Bacterial Viability ◽  
Binding Interface ◽  
Cryo Electron Microscopy ◽  
Stress Signals ◽  
Structural Insights

The stressosome transduces environmental stress signals to SigB to upregulate SigB-dependent transcription, which is required for bacterial viability. The stressosome core is composed of RsbS and at least one of the RsbR paralogs. A previous cryo-electron microscopy (cryo-EM) structure of the RsbRA–RsbS complex determined under a D2 symmetry restraint showed that the stressosome core forms a pseudo-icosahedron consisting of 60 STAS domains of RsbRA and RsbS. However, it is still unclear how RsbS and one of the RsbR paralogs assemble into the stressosome. Here, an assembly model of the stressosome is presented based on the crystal structure of the RsbS icosahedron and cryo-EM structures of the RsbRA–RsbS complex determined under diverse symmetry restraints (nonsymmetric C1, dihedral D2 and icosahedral I envelopes). 60 monomers of the crystal structure of RsbS fitted well into the I-restrained cryo-EM structure determined at 4.1 Å resolution, even though the STAS domains in the I envelope were averaged. This indicates that RsbS and RsbRA share a highly conserved STAS fold. 22 protrusions observed in the C1 envelope, corresponding to dimers of the RsbRA N-domain, allowed the STAS domains of RsbRA and RsbS to be distinguished in the stressosome core. Based on these, the model of the stressosome core was reconstructed. The mutation of RsbRA residues at the binding interface in the model (R189A/Q191A) significantly reduced the interaction between RsbRA and RsbS. These results suggest that nonconserved residues in the conserved STAS folds between RsbS and RsbR paralogs determine stressosome assembly.

scholarly journals Structural Insights into the BRAF Monomer-to-dimer Transition Mediated by RAS Binding

2021 ◽  
Author(s):  
Juliana Andrea Martinez Fiesco ◽  
David E Durrant ◽  
Deborah K Morrison ◽  
Ping Zhang
Keyword(s):  
Electron Microscopy ◽  
Conformational Changes ◽  
Dynamic Process ◽  
Mammalian Cells ◽  
Dual Function ◽  
Unresolved Issue ◽  
Raf Kinase ◽  
Cryo Electron Microscopy ◽  
Terminal Site ◽  
Structural Insights

An unresolved issue in RAF kinase signaling is how binding of autoinhibited RAF monomers to activated RAS initiates the conformational changes required to form active RAF dimers. Here, we present cryo-electron microscopy structures of full-length BRAF complexes derived from mammalian cells: autoinhibited monomeric BRAF:14-3-32:MEK and BRAF:14-3-32 complexes and an inhibitor-bound, dimeric BRAF2:14-3-32 complex, at 3.7, 4.1, and 3.9 Å resolution, respectively. The RAS binding domain (RBD) of BRAF is resolved in the autoinhibited structures, and we find that neither MEK nor ATP binding is required to stabilize the autoinhibited complexes. Notably, the RBD was found to interact extensively with the 14-3-3 protomer bound to the BRAF C-terminal site. Moreover, through structure-guided mutational studies, our findings indicate that RAS-RAF binding is a dynamic process and that RBD residues at the 14-3-3 interface have a dual function, first stabilizing RBD orientation in the autoinhibited state and then contributing to full RAS contact.

scholarly journals Structural Insights into TOR Signaling

Genes ◽  
2020 ◽  
Vol 11 (8) ◽  
pp. 885
Author(s):  
Lucas Tafur ◽  
Jennifer Kefauver ◽  
Robbie Loewith
Keyword(s):  
Electron Microscopy ◽  
Protein Kinase ◽  
Molecular Biology ◽  
Cellular Responses ◽  
Therapeutic Strategies ◽  
Cellular Growth ◽  
Tor Signaling ◽  
Cellular Homeostasis ◽  
Cryo Electron Microscopy ◽  
Structural Insights

The Target of Rapamycin (TOR) is a highly conserved serine/threonine protein kinase that performs essential roles in the control of cellular growth and metabolism. TOR acts in two distinct multiprotein complexes, TORC1 and TORC2 (mTORC1 and mTORC2 in humans), which maintain different aspects of cellular homeostasis and orchestrate the cellular responses to diverse environmental challenges. Interest in understanding TOR signaling is further motivated by observations that link aberrant TOR signaling to a variety of diseases, ranging from epilepsy to cancer. In the last few years, driven in large part by recent advances in cryo-electron microscopy, there has been an explosion of available structures of (m)TORC1 and its regulators, as well as several (m)TORC2 structures, derived from both yeast and mammals. In this review, we highlight and summarize the main findings from these reports and discuss both the fascinating and unexpected molecular biology revealed and how this knowledge will potentially contribute to new therapeutic strategies to manipulate signaling through these clinically relevant pathways.

scholarly journals Capsid structure of Leishmania RNA virus 1

2020 ◽  
Author(s):  
Michaela Procházková ◽  
Tibor Füzik ◽  
Danyil Grybchuk ◽  
Francesco Falginella ◽  
Lucie Podešvová ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Rational Design ◽  
Rna Binding ◽  
Rna Virus ◽  
Capsid Proteins ◽  
Viral Rnas ◽  
Mucocutaneous Leishmaniasis ◽  
Cryo Electron Microscopy ◽  
The Family ◽  
Positively Charged

Leishmania parasites cause a variety of symptoms, including mucocutaneous leishmaniasis, which results in the destruction of the mucous membranes of the nose, mouth, and throat. The species of Leishmania carrying Leishmania RNA virus 1 (LRV1), from the family Totiviridae, are more likely to cause severe disease and are less sensitive to treatment than those that do not contain the virus. Although the importance of LRV1 for the severity of leishmaniasis was discovered a long time ago, the structure of the virus remained unknown. Here, we present a cryo-electron microscopy reconstruction of the virus-like particle of LRV1 determined to a resolution of 3.65 Å. The capsid has icosahedral symmetry and is formed by 120 copies of a capsid protein assembled in asymmetric dimers. RNA genomes of viruses from the family Totiviridae are synthetized, but not capped at the 5’ end, by virus RNA-polymerases. To protect viral RNAs from degradation, capsid proteins of totivirus L-A cleave the 5’ caps of host mRNAs, creating decoys to overload the cellular RNA quality control system. Capsid proteins of LRV1 form positively charged clefts, which may be the cleavage sites for the 5’ cap of Leishmania mRNAs. Capsid proteins of LRV1 contain a putative RNA binding site distinct from that of the related L-A virus. The structure of the LRV1 capsid enables the rational design of compounds targeting the putative de-capping site. Such inhibitors may be developed into a treatment for mucocutaneous leishmaniasis caused by LRV1-positive species of Leishmania. IMPORTANCE Twelve million people worldwide suffer from leishmaniasis, resulting in more than thirty thousand deaths annually. The disease has several variants that differ in their symptoms. The mucocutaneous form, which leads to disintegration of the nasal septum, lips, and palate, is predominantly caused by Leishmania parasites carrying Leishmania RNA virus 1 (LRV1). Here, we present the structure of the LRV1 capsid determined using cryo-electron microscopy. Capsid proteins of a related totivirus L-A protect viral RNAs from degradation by cleaving the 5’ caps of host mRNAs. Capsid proteins of LRV1 may have the same function. We show that the LRV1 capsid contains positively charged clefts that may be sites for the cleavage of mRNAs of Leishmania cells. The structure of the LRV1 capsid enables the rational design of compounds targeting the putative mRNA cleavage site. Such inhibitors may be used as treatments for muco-cutaneous leishmaniasis.

scholarly journals Structural Insights Into the Initiation and Elongation of Ubiquitination by Ubr1

2021 ◽  
Author(s):  
Man Pan ◽  
Qingyun Zheng ◽  
Tian Wang ◽  
Lujun Liang ◽  
Junxiong Mao ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
E3 Ligase ◽  
Degradation Pathways ◽  
Single Chain ◽  
Cryo Electron Microscopy ◽  
N Terminus ◽  
Specific Manner ◽  
Ubiquitin Conjugating Enzyme ◽  
Structural Insights ◽  
Conjugating Enzyme

The N-end rule pathway was one of the first ubiquitin (Ub)-dependent degradation pathways to be identified. Ubr1, a single-chain E3 ligase, targets proteins bearing a destabilizing residue at the N-terminus (N-degron) for rapid K48-linked ubiquitination and proteasome-dependent degradation. How Ubr1 catalyses the initiation of ubiquitination on the substrate and elongation of the Ub chain in a linkage-specific manner through a single E2 ubiquitin-conjugating enzyme (Ubc2) remains unknown. Here, we report the cryo-electron microscopy structures of two complexes representing the initiation and elongation intermediates of Ubr1 captured using chemical approaches. In these two structures, Ubr1 adopts different conformations to facilitate the transfer of Ub from Ubc2 to either an N-degron peptide or a monoubiquitinated degron. These structures not only reveal the architecture of the Ubr1 complex but also provide mechanistic insights into the initiation and elongation steps of ubiquitination catalyzed by Ubr1.

scholarly journals Structural insights into the mechanism of human NPC1L1-mediated cholesterol uptake

2021 ◽  
Vol 7 (29) ◽  
pp. eabg3188
Author(s):  
Miaoqing Hu ◽  
Fan Yang ◽  
Yawen Huang ◽  
Xin You ◽  
Desheng Liu ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Cholesterol Level ◽  
Cholesterol Absorption ◽  
Bound State ◽  
Intestinal Cholesterol Absorption ◽  
Cholesterol Uptake ◽  
Cryo Electron Microscopy ◽  
Niemann Pick ◽  
Structural Insights ◽  
Apo State

Niemann-Pick C1-like 1 (NPC1L1) protein plays a central role in the intestinal cholesterol absorption and is the target of a drug, ezetimibe, which inhibits NPC1L1 to reduce cholesterol absorption. Here, we present cryo–electron microscopy structures of human NPC1L1 in apo state, cholesterol-enriched state, and ezetimibe-bound state to reveal molecular details of NPC1L1-mediated cholesterol uptake and ezetimibe inhibition. Comparison of these structures reveals that the sterol-sensing domain (SSD) could respond to the cholesterol level alteration by binding different number of cholesterol molecules. Upon increasing cholesterol level, SSD binds more cholesterol molecules, which, in turn, triggers the formation of a stable structural cluster in SSD, while binding of ezetimibe causes the deformation of the SSD and destroys the structural cluster, leading to the inhibition of NPC1L1 function. These results provide insights into mechanisms of NPC1L1 function and ezetimibe action and are of great significance for the development of new cholesterol absorption inhibitors.

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