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Herpesviridae comprise a large family of enveloped DNA viruses all of whom employ orthologs of the same three glycoproteins, gB, gH and gL. Additionally, herpesviruses often employ accessory proteins to bind receptors and/or bind ...
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Herpesviridae comprise a large family of enveloped DNA viruses all of whom employ orthologs of the same three glycoproteins, gB, gH and gL. Additionally, herpesviruses often employ accessory proteins to bind receptors and/or bind the heterodimer gH/gL or even to determine cell tropism. Sorting out how these proteins function has been resolved to a large extent by structural biology coupled with supporting biochemical and biologic evidence. Together with the G protein of vesicular stomatitis virus, gB is a charter member of the Class III fusion proteins. Unlike VSV G, gB only functions when partnered with gH/gL. However, gH/gL does not resemble any known viral fusion protein and there is evidence that its function is to upregulate the fusogenic activity of gB. In the case of herpes simplex virus, gH/gL itself is upregulated into an active state by the conformational change that occurs when gD, the receptor binding protein, binds one of its receptors. In this review we focus primarily on prototypes of the three subfamilies of herpesviruses. We will present our model for how herpes simplex virus (HSV) regulates fusion in series of highly regulated steps. Our model highlights what is known and also provides a framework to address mechanistic questions about fusion by HSV and herpesviruses in general.
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Negative-stain transmission electron microscopy (EM) is a technique that hasprovided nanometer resolution images of macromolecules for about 60 years.Developments in cryo-EM image processing have maximized the informationgained fr...
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Negative-stain transmission electron microscopy (EM) is a technique that hasprovided nanometer resolution images of macromolecules for about 60 years.Developments in cryo-EM image processing have maximized the informationgained from averaging large numbers of particles. These developments cannow be applied back to negative-stain image analysis to ascertain domain levelmolecular structure (10 to 20 A) more quickly and efficiently than possibleby atomic resolution cryo-EM. Using uranyl acetate stained molecular complexesof influenza hemagglutinin bound to Fab 441D6, we describe a simpleand efficient means to collect several hundred micrographs with SerialEM.Using RELION, we illustrate how tens of thousands of complexes can beauto-picked and classified to accurately describe the domain level topologyof this unconventional hemagglutinin head-domain epitope. By comparing tothe cryo-EM density map of the same complex, we show that questions aboutepitope mapping and conformational heterogeneity can readily be answered bythis negative-stain method.
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This article advocates for web scraping as an effective method to augment and enhance technical and professional communication (TPC) research practices. Web scraping is used to create consistently structured and well-sampled data ...
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This article advocates for web scraping as an effective method to augment and enhance technical and professional communication (TPC) research practices. Web scraping is used to create consistently structured and well-sampled data sets about domains, communities, demographics, and topics of interest to TPC scholars. After providing an extended description of web scraping, the authors identify technical considerations of the method and provide practitioner narratives. They then describe an overview of project-oriented web scraping. Finally, they discuss implications for the concept as a sustainable approach to developing web scraping methods for TPC research.
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This article offers a methodology for conducting large-scale audience analysis called "big data audience analysis" (BDAA). BDAA uses distant reading and thin description to examine a large corpus of text data from online audiences...
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This article offers a methodology for conducting large-scale audience analysis called "big data audience analysis" (BDAA). BDAA uses distant reading and thin description to examine a large corpus of text data from online audiences. In this article, that corpus is approximately 450,000 online reader comments. We analyze this corpus through sentiment analysis, statistical analysis, and geolocation to identify trends and patterns in large datasets. BDAA can better prepare TPC researchers for large-scale audience studies.
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Received 11 May 2015. Accepted 18 June 2015. Accepted manuscript posted online 24 June 2015. Address correspondence to Tina M. Cairns, tmcairns{at}dental.upenn.edu. T.M.C. and Z.-Y.H. contributed equally to this article. Citation ...
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Received 11 May 2015. Accepted 18 June 2015. Accepted manuscript posted online 24 June 2015. Address correspondence to Tina M. Cairns, tmcairns{at}dental.upenn.edu. T.M.C. and Z.-Y.H. contributed equally to this article. Citation Cairns TM, Huang Z-Y, Gallagher JR, Lin Y, Lou H, Whitbeck JC, Wald A, Cohen GH, Eisenberg RJ. 2015. Patient-specific neutralizing antibody responses to herpes simplex virus are attributed to epitopes on gD, gB, or both and can be type specific. J Virol 89:9213–9231. doi:10.1128/JVI.01213-15.
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Immunoelectron microscopy is a powerful technique for identifying viral antigensand determining their structural localization and organization within vaccinesand viruses. While traditional negative staining transmission electronmi...
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Immunoelectron microscopy is a powerful technique for identifying viral antigensand determining their structural localization and organization within vaccinesand viruses. While traditional negative staining transmission electronmicroscopy provides structural information, identity of components within asample may be confounding. Immunoelectron microscopy allows for identifi-cation and visualization of antigens and their relative positions within a particulatesample. This allows for simple qualitative analysis of samples includingwhole virus, viral components, and viral-like particles. This article describesmethods for immunogold labeling of viral antigens in a liquid suspension, withexamples of immunogold-labeled influenza virus glycoproteins, and also discussesthe important considerations for sample preparation and determinationof morphologies. Together, these methods allow for understanding the antigenicmakeup of viral particulate samples, which have important implicationsfor molecular virology and vaccine development. C 2019 The Authors. Thisis an open access article under the terms of the Creative Commons AttributionLicense, which permits use, distribution and reproduction in any medium,provided the original work is properly cited.
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ABSTRACT All enveloped viruses, including herpesviruses, must fuse their envelope with the host membrane to deliver their genomes into target cells, making this essential step subject to interference by antibodies and drugs. Viral...
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ABSTRACT All enveloped viruses, including herpesviruses, must fuse their envelope with the host membrane to deliver their genomes into target cells, making this essential step subject to interference by antibodies and drugs. Viral fusion is mediated by a viral surface protein that transits from an initial prefusion conformation to a final postfusion conformation. Strikingly, the prefusion conformation of the herpesvirus fusion protein, gB, is poorly understood. Herpes simplex virus (HSV), a model system for herpesviruses, causes diseases ranging from mild skin lesions to serious encephalitis and neonatal infections. Using cryo-electron tomography and subtomogram averaging, we have characterized the structure of the prefusion conformation and fusion intermediates of HSV-1 gB. To this end, we have set up a system that generates microvesicles displaying full-length gB on their envelope. We confirmed proper folding of gB by nondenaturing electrophoresis-Western blotting with a panel of monoclonal antibodies (MAbs) covering all gB domains. To elucidate the arrangement of gB domains, we labeled them by using (i) mutagenesis to insert fluorescent proteins at specific positions, (ii) coexpression of gB with Fabs for a neutralizing MAb with known binding sites, and (iii) incubation of gB with an antibody directed against the fusion loops. Our results show that gB starts in a compact prefusion conformation with the fusion loops pointing toward the viral membrane and suggest, for the first time, a model for gB’s conformational rearrangements during fusion. These experiments further illustrate how neutralizing antibodies can interfere with the essential gB structural transitions that mediate viral entry and therefore infectivity. IMPORTANCE The herpesvirus family includes herpes simplex virus (HSV) and other human viruses that cause lifelong infections and a variety of diseases, like skin lesions, encephalitis, and cancers. As enveloped viruses, herpesviruses must fuse their envelope with the host membrane to start an infection. This process is mediated by a viral surface protein that transitions from an initial conformation (prefusion) to a final, more stable, conformation (postfusion). However, the prefusion conformation of the herpesvirus fusion protein (gB) is poorly understood. To elucidate the structure of the prefusion conformation of HSV type 1 gB, we have employed cryo-electron microscopy to study gB molecules expressed on the surface of vesicles. Using different approaches to label gB’s domains allowed us to model the structures of the prefusion and intermediate conformations of gB. Overall, our findings enhance our understanding of HSV fusion and lay the groundwork for the development of new ways to prevent and block HSV infection.
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Providing contextualized, effective writing instruction for engineering students is an important and challenging objective. This article presents a needs analysis conducted in a large engineering college and introduces the faculty...
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Providing contextualized, effective writing instruction for engineering students is an important and challenging objective. This article presents a needs analysis conducted in a large engineering college and introduces the faculty development program that was created based on that analysis. The authors advocate for sustained interdisciplinary collaboration to promote contextualized adoption and adaptation of best practices and testing of scalable strategies.
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Influenza virus infects millions of people annually and can cause global pandemics. Hemagglutinin (HA) is the primary component of commercial influenza vaccines (CIV), and antibody titer to HA is a primary correlate of protection....
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Influenza virus infects millions of people annually and can cause global pandemics. Hemagglutinin (HA) is the primary component of commercial influenza vaccines (CIV), and antibody titer to HA is a primary correlate of protection. Continual antigenic variation of HA requires that CIVs are reformulated yearly. Structural organization of HA complexes have not previously been correlated with induction of broadly reactive antibodies, yet CIV formulations vary in how HA is organized. Using electron microscopy to study four current CIVs, we find structures including: individual HAs, starfish structures with up to 12 HA molecules, and novel spiked-nanodisc structures that display over 50 HA molecules along the complex’s perimeter. CIV containing these spiked nanodiscs elicit the highest levels of heterosubtypic cross-reactive antibodies in female mice. Here, we report that HA structural organization can be an important CIV parameter and can be associated with the induction of cross-reactive antibodies to conserved HA epitopes. Here, Myers and Gallagher et al. characterize the structural organization of commercial influenza vaccines. The vaccines differ in their structural composition and identify a “spiked nanodisc” arrangement of hemagglutinin (HA) with increased display and immunogenicity of the conserved stem region of HA.
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Prompted by an in-depth case study of a web-writer, this article argues that audience may be understood as an emergent process for web-writers who consider their comments. Rather than a group of people or demographic that is prefi...
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Prompted by an in-depth case study of a web-writer, this article argues that audience may be understood as an emergent process for web-writers who consider their comments. Rather than a group of people or demographic that is prefigured to exist, this article posits that audience might be a concept used throughout the composing process, including production and distribution processes. Such an approach is useful for digital pedagogies that involve online comments because it allows audience to be considered before, during, and after writers' production processes.
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