Michael G Rossmann

Purdue University, ウェストラファイエット, Indiana, United States

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Publications (511)3236.22 Total impact

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    ABSTRACT: Enterovirus D68 (EV-D68) is an emerging pathogen that can cause severe respiratory disease and is associated with cases of paralysis, especially among children. Heretofore, information on host factor requirements for EV-D68 infection is scarce. Haploid genetic screening is a powerful tool to reveal factors involved in the entry of pathogens. We performed a genome-wide haploid screen with the EV-D68 prototype Fermon strain to obtain a comprehensive overview of cellular factors supporting EV-D68 infection. We identified and confirmed several genes involved in sialic acid (Sia) biosynthesis, transport, and conjugation to be essential for infection. Moreover, by using knockout cell lines and gene reconstitution, we showed that both α2,6- and α2,3-linked Sia can be used as functional cellular EV-D68 receptors. Importantly, the screen did not reveal a specific protein receptor, suggesting that EV-D68 can use multiple redundant sialylated receptors. Upon testing recent clinical strains, we identified strains that showed a similar Sia dependency, whereas others could infect cells lacking surface Sia, indicating they can use an alternative, nonsialylated receptor. Nevertheless, these Sia-independent strains were still able to bind Sia on human erythrocytes, raising the possibility that these viruses can use multiple receptors. Sequence comparison of Sia-dependent and Sia-independent EV-D68 strains showed that many changes occurred near the canyon that might allow alternative receptor binding. Collectively, our findings provide insights into the identity of the EV-D68 receptor and suggest the possible existence of Sia-independent viruses, which are essential for understanding tropism and disease.
    Full-text · Article · Jan 2016 · Proceedings of the National Academy of Sciences
  • Xinzheng Zhang · Michael G. Rossmann

    No preview · Article · Dec 2015 · Current Opinion in Virology
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    ABSTRACT: Human enterovirus D68 (EV-D68) is a causative agent of childhood respiratory diseases and has now emerged as a global public health threat. Nevertheless, knowledge of the tissue tropism and pathogenesis of EV-D68 has been hindered by a lack of studies on the receptor-mediated EV-D68 entry into host cells. Here we demonstrate that cell surface sialic acid is essential for EV-D68 to bind to and infect susceptible cells. Crystal structures of EV-D68 in complex with sialylated glycan receptor analogues show that they bind into the 'canyon' on the virus surface. The sialic acid receptor induces a cascade of conformational changes in the virus to eject a fatty-acid-like molecule that regulates the stability of the virus. Thus, virus binding to a sialic acid receptor and to immunoglobulin-like receptors used by most other enteroviruses share a conserved mechanism for priming viral uncoating and facilitating cell entry.
    Full-text · Article · Nov 2015 · Nature Communications
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    ABSTRACT: We screened a panel of mouse and human monoclonal antibodies (MAbs) against chikungunya virus and identified several with inhibitory activity against multiple alphaviruses. Passive transfer of broadly neutralizing MAbs protected mice against infection by chikungunya, Mayaro, and O'nyong'nyong alphaviruses. Using alanine-scanning mutagenesis, loss-of-function recombinant proteins and viruses, and multiple functional assays, we determined that broadly neutralizing MAbs block multiple steps in the viral lifecycle, including entry and egress, and bind to a conserved epitope on the B domain of the E2 glycoprotein. A 16 Å resolution cryo-electron microscopy structure of a Fab fragment bound to CHIKV E2 B domain provided an explanation for its neutralizing activity. Binding to the B domain was associated with repositioning of the A domain of E2 that enabled cross-linking of neighboring spikes. Our results suggest that B domain antigenic determinants could be targeted for vaccine or antibody therapeutic development against multiple alphaviruses of global concern.
    No preview · Article · Nov 2015 · Cell
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    ABSTRACT: Chikungunya is a positive-stranded RNA alphavirus. Structures of chikungunya virus-like particles in complex with strongly neutralizing antibody Fab fragments (8B10 and 5F10) were determined using cryo-electron microscopy and X-ray crystallography. By fitting the crystallographically determined structures of these Fab fragments into the cryo-electron density maps, we show that Fab fragments of antibody 8B10 extend radially from the viral surface and block receptor-binding on the E2 glycoprotein. In contrast, Fab fragments of antibody 5F10 bind the tip of the E2 B domain and lie tangentially on the viral surface. Fab 5F10 fixes the B domain rigidly to the surface of the virus, blocking exposure of the fusion loop on glycoprotein E1, and therefore preventing the virus from becoming fusogenic. Although Fab 5F10 can neutralize the wild type virus, it can also bind to a mutant virus without inhibiting fusion or attachment. Although the mutant virus is no longer able to propagate by extracellular budding, it can however enter the next cell by travelling through junctional complexes without being intercepted by a neutralizing antibody to the wild-type virus, thus clarifying how cell-to-cell transmission can occur. IMPORTANCE Alphaviral infections are mainly transmitted by mosquitoes. Chikungunya virus (CHIKV), which belongs to the alphavirus genus, has a wide distribution in the Old World that has expanded in recent years into the Americas. There are currently no vaccines or drugs against alphaviral infections. Therefore, a better understanding of CHIKV and its associated neutralizing antibodies will aid in the development of effective treatments.
    No preview · Article · Nov 2015 · Journal of Virology
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    ABSTRACT: Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes severe acute and chronic disease in humans. Although highly inhibitory murine and human monoclonal antibodies (mAbs) have been generated, the structural basis of their neutralizing activity remains poorly characterized. Here, we determined the cryo-EM structures of chikungunya virus-like particles complexed with antibody fragments (Fab) of two highly protective human mAbs, 4J21 and 5M16, that block virus fusion with host membranes. Both mAbs bind primarily to sites within the A and B domains, as well as to the B domain's β-ribbon connector of the viral glycoprotein E2. The footprints of these antibodies on the viral surface were consistent with results from loss-of-binding studies using an alanine scanning mutagenesis-based epitope mapping approach. The Fab fragments stabilized the position of the B domain relative to the virus, particularly for the complex with 5M16. This finding is consistent with a mechanism of neutralization in which anti-CHIKV mAbs that bridge the A and B domains impede movement of the B domain away from the underlying fusion loop on the E1 glycoprotein and therefore block the requisite pH-dependent fusion of viral and host membranes.
    No preview · Article · Oct 2015 · Proceedings of the National Academy of Sciences
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    ABSTRACT: Epstein-Barr virus (EBV) represents a major global health problem. Though it is associated with infectious mononucleosis and ∼200,000 cancers annually worldwide, a vaccine is not available. The major target of immunity is EBV glycoprotein 350/220 (gp350) that mediates attachment to B cells through complement receptor 2 (CR2/CD21). Here, we created self-assembling nanoparticles that displayed different domains of gp350 in a symmetric array. By focusing presentation of the CR2-binding domain on nanoparticles, potent neutralizing antibodies were elicited in mice and non-human primates. The structurally designed nanoparticle vaccine increased neutralization 10- to 100-fold compared to soluble gp350 by targeting a functionally conserved site of vulnerability, improving vaccine-induced protection in a mouse model. This rational approach to EBV vaccine design elicited potent neutralizing antibody responses by arrayed presentation of a conserved viral entry domain, a strategy that can be applied to other viruses. Copyright © 2015 Elsevier Inc. All rights reserved. Full text can be found at http://authors.elsevier.com/a/1RcBdL7PXI6a- Valid until October 16, 2015
    No preview · Article · Aug 2015 · Cell
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    ABSTRACT: The structure and assembly of bacteriophage T4 has been extensively studied. However, the detailed structure of the portal protein remained unknown. Here we report the structure of the bacteriophage T4 portal assembly, gene product 20 (gp20), determined by cryo-electron microscopy (cryo-EM) to 3.6 Å resolution. In addition, analysis of a 10 Å resolution cryo-EM map of an empty prolate T4 head shows how the dodecameric portal assembly interacts with the capsid protein gp23 at the special pentameric vertex. The gp20 structure also verifies that the portal assembly is required for initiating head assembly, for attachment of the packaging motor, and for participation in DNA packaging. Comparison of the Myoviridae T4 portal structure with the known portal structures of φ29, SPP1 and P22, representing Podo- and Siphoviridae, shows that the portal structure probably dates back to a time when self-replicating microorganisms were being established on Earth.
    Full-text · Article · Jul 2015 · Nature Communications
  • Xinzheng Zhang · Lei Sun · Michael G Rossmann
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    ABSTRACT: Dengue virus is the causative agent of dengue virus fever. It infects about 400 million people per year and leads to about 21,000 deaths annually. There is available neither a fully successful vaccine nor a successful drug therapy. Some dengue virus serotypes undergo a temperature dependent conformational change from a 'smooth' form at lower temperatures to a 'bumpy' form at temperatures approaching 37°C, the human body temperature. The bumpy structure is less stable and is probably an intermediate in the formation of a fusogenic virus particle. Copyright © 2015 Elsevier B.V. All rights reserved.
    No preview · Article · Jun 2015 · Current Opinion in Virology
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    ABSTRACT: Mimivirus was initially identified as a bacterium because its dense, 125-nm-long fibers stained Gram-positively. These fibers probably play a role during the infection of some host cells. The normal hosts of Mimivirus are unknown, but in the laboratory Mimivirus is usually propagated in amoeba. The structure of R135, a major component of the fibrous outer layer of Mimivirus, has been determined to 2-Å resolution. The protein's structure is similar to that of members of the glucose-methanol-choline oxidoreductase family, which have an N-terminal FAD binding domain and a C-terminal substrate recognition domain. The closest homolog to R135 is an aryl-alcohol oxidase that participates in lignin biodegradation of plant cell walls. Thus R135 might participate in the degradation of their normal hosts, including some lignin-containing algae. Copyright © 2015 Elsevier Ltd. All rights reserved.
    Full-text · Article · Apr 2015 · Structure
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    ABSTRACT: Giant viruses are protist-associated viruses belonging to the proposed order Megavirales; almost all have been isolated on Acanthamoeba sp. Their isolation in humans suggests that they are part of the human virome. Using a high-throughput strategy to isolate new giant viruses on their original protozoa hosts, we obtained eight isolates of a new giant viral lineage on Vermamoeba vermiformis, the most common free-living protist found in human environments. This new lineage was proposed to be a Faustovirus. The prototype member Faustovirus E12 forms icosahedral virions of ≈200 nm in size that are devoid of fibrils and that encapsidate a 466 kilobase pair-long genome encoding 451 predicted proteins. Of these, 164 are found in the virion. The phylogenetic analysis of core viral genes shows that Faustovirus is distantly related to the mammalian pathogen African swine fever virus, but encodes ≈3 times more mosaic gene complements. About two-thirds of these genes do not show significant similarity to any known proteins. These findings show that expanding the panel of protists to discover new giant viruses is a fruitful strategy. By using for the first time Vermamoeba, a protist living in humans and their environment, we isolated eight strains of a new giant virus we named Faustovirus. The genomes of these strains were sequenced, showing that the faustoviruses are related but different from the vertebrate pathogen African swine fever virus (ASFV), which belongs to the family Asfarviridae. Moreover, the Faustovirus gene repertoire is ≈3 times larger than that of ASFV and comprises ≈ two-thirds ORFans. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Full-text · Article · Apr 2015 · Journal of Virology
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    ABSTRACT: Enterovirus D68 (EV-D68) is a member of Picornaviridae and is a causative agent of recent outbreaks of respiratory illness in children in the United States. We report here the crystal structures of EV-D68 and its complex with pleconaril, a capsid-binding compound that had been developed as an anti-rhinovirus drug. The hydrophobic drug-binding pocket in viral protein 1 contained density that is consistent with a fatty acid of about 10 carbon atoms. This density could be displaced by pleconaril. We also showed that pleconaril inhibits EV-D68 at a half-maximal effective concentration of 430 nanomolar and might, therefore, be a possible drug candidate to alleviate EV-D68 outbreaks. Copyright © 2015, American Association for the Advancement of Science.
    No preview · Article · Jan 2015 · Science
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    ABSTRACT: Unlabelled: Flaviviruses undergo large conformational changes during their life cycle. Under acidic pH conditions, the mature virus forms transient fusogenic trimers of E glycoproteins that engage the lipid membrane in host cells to initiate viral fusion and nucleocapsid penetration into the cytoplasm. However, the dynamic nature of the fusogenic trimer has made the determination of its structure a challenge. Here we have used Fab fragments of the neutralizing antibody DV2-E104 to stop the conformational change of dengue virus at an intermediate stage of the fusion process. Using cryo-electron microscopy, we show that in this intermediate stage, the E glycoproteins form 60 trimers that are similar to the predicted "open" fusogenic trimer. Importance: The structure of a dengue virus has been captured during the formation of fusogenic trimers. This was accomplished by binding Fab fragments of the neutralizing antibody DV2-E104 to the virus at neutral pH and then decreasing the pH to 5.5. These trimers had an "open" conformation, which is distinct from the "closed" conformation of postfusion trimers. Only two of the three E proteins within each spike are bound by a Fab molecule at domain III. Steric hindrance around the icosahedral 3-fold axes prevents binding of a Fab to the third domain III of each E protein spike. Binding of the DV2-E104 Fab fragments prevents domain III from rotating by about 130° to the postfusion orientation and thus precludes the stem region from "zipping" together the three E proteins along the domain II boundaries into the "closed" postfusion conformation, thus inhibiting fusion.
    Full-text · Article · Oct 2014 · Journal of Virology
  • Moh Lan Yap · Michael G Rossmann
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    ABSTRACT: ABSTRACT Bacteriophage T4 is the most well-studied member of Myoviridae, the most complex family of tailed phages. T4 assembly is divided into three independent pathways: the head, the tail and the long tail fibers. The prolate head encapsidates a 172 kbp concatemeric dsDNA genome. The 925 Å-long tail is surrounded by the contractile sheath and ends with a hexagonal baseplate. Six long tail fibers are attached to the baseplate's periphery and are the host cell's recognition sensors. The sheath and the baseplate undergo large conformational changes during infection. X-ray crystallography and cryo-electron microscopy have provided structural information on protein-protein and protein-nucleic acid interactions that regulate conformational changes during assembly and infection of Escherichia coli cells.
    No preview · Article · Oct 2014 · Future Microbiology
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    ABSTRACT: Chloroviruses infect their hosts by specifically binding to and degrading the cell wall of their algal hosts at the site of attachment, using an intrinsic digesting enzyme(s). Chlorovirus PBCV-1 stored as a lysate survived longer than virus alone, suggesting virus attachment to cellular debris may be reversible. Ghost cells (algal cells extracted with methanol) were used as a model to study reversibility of PBCV-1 attachment because ghost cells are as susceptible to attachment and wall digestion as are live cells. Reversibility of attachment to ghost cells was examined by releasing attached virions with a cell wall degrading enzyme extract. The majority of the released virions retained infectivity even after re-incubating the released virions with ghost cells two times. Thus the chloroviruses appear to have a dynamic attachment strategy that may be beneficial in indigenous environments where cell wall debris can act as a refuge until appropriate host cells are available.
    Full-text · Article · Sep 2014 · Virology
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    Michael G Rossmann
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    ABSTRACT: I describe my gradually evolving role as a scientist from my birth in Frankfurt (Germany) to my education in the UK, my post-doc years and my experiences as an independent investigator at Purdue University1. I discuss the significance of my post-doctoral work in Minnesota where I had my first encounter with an electronic computer and subsequently in Cambridge where I participated in the first structure determination of proteins. After six years back in England my family moved to Indiana (USA) where my home remains to this day. At Purdue University I first studied the structure of enzymes and in the process I discovered the organization and slow evolution of protein domains, each with a specific function. With this success I started what had been on my mind already for a long time, namely the structural analysis of viruses. Initially we studied plant viruses but then switched to small RNA animal viruses, discovering that some plant and animal RNA viruses have closely similar structures and therefore presumably had a common evolutionary origin. Next I became interested in somewhat larger viruses that had lipid membrane envelopes. In turn that has led to the study of very large dsDNA viruses as big as small bacteria as well as studies of bacterial viruses that require complex molecular motors for different parts of their life cycle. While developing crystallographic techniques for the study of viruses it has become progressively more apparent that electron microscopy is an important new tool that is likely to eclipse x-ray crystallography in the next decade.
    Preview · Article · Sep 2014 · Physica Scripta
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    Yue Liu · Michael G Rossmann

    Preview · Article · Aug 2014 · Protein & Cell
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    ABSTRACT: PhiX174 is a small circular ssDNA icosahedral phage of known structure (McKenna et al., 1992). The infectious viral capsid consists of 60 copies of the major capsid protein F, 60 copies of the spike protein G, 60 copies of the DNA binding protein J as well as about 12 copies of the DNA pilot protein H. We have shown that the H protein can assemble in vitro and in vivo into tubes that are wide enough to allow the passage of two antiparallel strands of ssDNA. We will give details of our mutational and cryo electron tomographic analyses showing that these tubes are required for the translocation of the viral genome into the cytoplasm of the E.Coli host. Thus the mechanism of infection of tail-less phages like phiX174 and of tailed phages may be a case of convergent evolution.
    Full-text · Article · Aug 2014 · Acta Crystallographica Section A: Foundations and Advances
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    ABSTRACT: The bacteriophage T4 baseplate is the control center of the virus, where the recognition of an E. coli host by the long tail fibers is translated into a signal to initiate infection. The short tail fibers unfold from the baseplate for firm attachment to the host, followed by shrinkage of the tail sheath that causes the tail tube to enter and cross the periplasmic space ending with injection of the genome into the host. During this process, the 6.5 MDa baseplate changes its structure from a "dome" shape to a "star" shape. An in vitro assembled hubless baseplate has been crystalized. It consists of six copies of the recombinantly expressed trimeric gene product (gp) 10, monomeric gp7, dimeric gp8, dimeric gp6 and monomeric gp53. The diffraction pattern extends, at most, to 4.0 Å resolution. The known partial structures of gp10, gp8, and gp6 and their relative position in the baseplate derived from earlier electron microscopy studies were used for molecular replacement. An electron density map has been calculated based on molecular replacement, single isomorphous replacement with anomalous dispersion data and 2-fold non-crystallographic symmetry averaging between two baseplate wedges in the crystallographic asymmetric unit. The current electron density map indicates that there are structural changes in the gp6, gp8, and gp10 oligomers compared to their structures when separately crystallized. Additional density is also visible corresponding to gp7, gp53 and the unknown parts of gp10 and gp6.
    No preview · Article · Jul 2014 · Journal of Structural Biology
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    Lei Sun · Michael G Rossmann · Bentley A Fane
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    ABSTRACT: Although ϕX174 DNA pilot protein H is monomeric during procapsid assembly, it forms an oligomeric tube on the host cell surface. Reminiscent of a double-stranded DNA phage tail in form and function, the H tube transports the single-stranded ϕX174 genome across the Escherichia coli cell wall. The 2.4-Å resolution H-tube crystal structure suggests functional and energetic mechanisms that may be common features of DNA transport through virally encoded conduits.
    Full-text · Article · Jul 2014 · Journal of Virology

Publication Stats

31k Citations
3,236.22 Total Impact Points

Institutions

  • 1966-2015
    • Purdue University
      • Department of Biological Sciences
      ウェストラファイエット, Indiana, United States
  • 2011
    • Columbia University
      New York City, New York, United States
    • Chiang Mai University
      • Department of Microbiology
      Amphoe Muang Chiang Mai, Chiang Mai, Thailand
  • 2004-2011
    • The Catholic University of America
      • Department of Biology
      Washington, Washington, D.C., United States
  • 2010
    • National Institute of Arthritis and Musculoskeletal and Skin Diseases
      Maryland, United States
    • Southern Research Institute
      Birmingham, Alabama, United States
  • 2009
    • French National Centre for Scientific Research
      • Laboratoire Information Génomique et Structurale (IGS)
      Lutetia Parisorum, Île-de-France, France
  • 1994-2009
    • Cornell University
      • College of Veterinary Medicine
      Ithaca, New York, United States
  • 2008
    • Case Western Reserve University
      • Institute of Pathology
      Cleveland, Ohio, United States
    • Universität Regensburg
      • Department of Medical Microbiology and Hygiene
      Ratisbon, Bavaria, Germany
  • 2007
    • Universidad Autónoma de Madrid
      Madrid, Madrid, Spain
  • 2000
    • The University of Manchester
      Manchester, England, United Kingdom
  • 1995
    • Florida State University
      • Institute of Molecular Biophysics
      Tallahassee, Florida, United States
  • 1993
    • Justus-Liebig-Universität Gießen
      • Institut für Virologie
      Gießen, Hesse, Germany
  • 1985-1989
    • University of Wisconsin–Madison
      • Institute for Molecular Virology
      Madison, Wisconsin, United States
  • 1983
    • Uppsala University
      • Department of Cell and Molecular Biology
      Uppsala, Uppsala, Sweden
  • 1982
    • Mrc Harwell
      Oxford, England, United Kingdom