scholarly journals Bunyamwera virus possesses a distinct nucleocapsid protein to facilitate genome encapsidation

2013 ◽  
Vol 110 (22) ◽  
pp. 9048-9053 ◽  
Author(s):  
B. Li ◽  
Q. Wang ◽  
X. Pan ◽  
I. Fernandez de Castro ◽  
Y. Sun ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Genome Encapsidation ◽  
Bunyamwera Virus

scholarly journals Homotypic Interaction of Bunyamwera Virus Nucleocapsid Protein

2005 ◽  
Vol 79 (20) ◽  
pp. 13166-13172 ◽  
Author(s):  
Vincent H. J. Leonard ◽  
Alain Kohl ◽  
Jane C. Osborne ◽  
Angela McLees ◽  
Richard M. Elliott
Keyword(s):  
Nucleocapsid Protein ◽  
Cross Linking ◽  
Genomic Rna ◽  
Rna Dependent Rna Polymerase ◽  
N Protein ◽  
Two Hybrid ◽  
Chemical Cross Linking ◽  
Bunyamwera Virus ◽  
Transcription And Replication ◽  
Weight Structures

ABSTRACT The bunyavirus nucleocapsid protein, N, plays a central role in viral replication in encapsidating the three genomic RNA segments to form functional templates for transcription and replication by the viral RNA-dependent RNA polymerase. Here we report functional mapping of interacting domains of the Bunyamwera orthobunyavirus N protein by yeast and mammalian two-hybrid systems, immunoprecipitation experiments, and chemical cross-linking studies. N forms a range of multimers from dimers to high-molecular-weight structures, independently of the presence of RNA. Deletion of the N- or C-terminal domains resulted in loss of activity in a minireplicon assay and a decreased capacity for N to form higher multimers. Our data suggest a head-to-head and tail-to-tail multimerization model for the orthobunyavirus N protein.

scholarly journals RNA Binding Properties of Bunyamwera Virus Nucleocapsid Protein and Selective Binding to an Element in the 5′ Terminus of the Negative-Sense S Segment

2000 ◽  
Vol 74 (21) ◽  
pp. 9946-9952 ◽  
Author(s):  
Jane C. Osborne ◽  
Richard M. Elliott
Keyword(s):  
Nucleocapsid Protein ◽  
Rna Binding ◽  
Metal Chelate ◽  
Rnase A ◽  
Mobility Shift ◽  
N Protein ◽  
Encapsidation Signal ◽  
Negative Sense ◽  
Bunyamwera Virus

ABSTRACT The genome of Bunyamwera virus (BUN) (familyBunyaviridae, genus Bunyavirus) comprises three negative-sense RNA segments which act as transcriptional templates for the viral polymerase only when encapsidated by the nucleocapsid protein (N). Previous studies have suggested that the encapsidation signal may reside within the 5′ terminus of each segment. The BUN N protein was expressed as a 6-histidine-tagged fusion protein in Escherichia coli and purified by metal chelate chromatography. An RNA probe containing the 5′-terminal 32 and 3′-terminal 33 bases of the BUN S (small) genome segment was used to investigate binding by the N protein in vitro using gel mobility shift and filter binding assays. On acrylamide gels a number of discrete RNA-N complexes were resolved, and analysis of filter binding data indicated a degree of cooperativity in N protein binding. RNA-N complexes were resistant to digestion with up to 1 μg of RNase A per ml. Competition assays with a variety of viral and nonviral RNAs identified a region within the 5′ terminus of the BUN S segment for which N had a high preference for binding. This site may constitute the signal for initiation of encapsidation by N.

scholarly journals The structure of a native orthobunyavirus ribonucleoprotein reveals a key role for viral RNA in maintaining its helical architecture

2021 ◽  
Author(s):  
Francis Hopkins ◽  
Beatriz Alvarez-Rodriguez ◽  
George Heath ◽  
Kyriakoulla Panayi ◽  
Samantha Hover ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Rna Synthesis ◽  
Rna Replication ◽  
Longitudinal Axis ◽  
Emerging Pathogens ◽  
Cryo Electron Microscopy ◽  
Therapeutic Measures ◽  
Human Use ◽  
Negative Sense ◽  
Bunyamwera Virus

The Bunyavirales order of RNA viruses comprises emerging pathogens for which approved preventative or therapeutic measures for human use are not available. The genome of all Bunyavirales consists of negative-sense RNA segments wrapped by the virus-encoded nucleocapsid protein (NP) to form ribonucleoproteins (RNPs). RNPs represent the active template for RNA synthesis and the form in which the genome is packaged into virions, functions that require inherent flexibility. We present a pseudo-atomic model of a native RNP purified from Bunyamwera virus (BUNV), the prototypical Bunyavirales member, based on a cryo-electron microscopy (cryo-EM) average at 13 A resolution with subsequent fitting of the BUNV NP crystal structure by molecular dynamics. We show the BUNV RNP possesses relaxed helical architecture, with successive helical turns separated by ~18 A. The model shows that adjacent NP monomers in the RNP chain interact laterally through flexible N- and C-terminal arms, with no helix-stabilizing interactions along the longitudinal axis. Instead, EM analysis of RNase-treated RNPs suggests their chain integrity is dependent on the encapsidated genomic RNA, thus providing the molecular basis for RNP flexibility. Overall, this work will assist in designing anti-viral compounds targeting the RNP and inform studies on bunyaviral RNP assembly, packaging and RNA replication.

scholarly journals Elucidation of functional domains of Chandipura virus Nucleocapsid protein involved in oligomerization and RNA binding: Implication in viral genome encapsidation

Virology ◽  
2010 ◽  
Vol 407 (1) ◽  
pp. 33-42 ◽  
Author(s):  
Arindam Mondal ◽  
Raja Bhattacharya ◽  
Tridib Ganguly ◽  
Subhradip Mukhopadhyay ◽  
Atanu Basu ◽  
...  
Keyword(s):  
Viral Genome ◽  
Nucleocapsid Protein ◽  
Rna Binding ◽  
Functional Domains ◽  
Chandipura Virus ◽  
Genome Encapsidation

Antibody Screening Results for Anti-Nucleocapsid Antibodies Towards the Development of a SARS-CoV-2 Nucleocapsid Protein Antigen Detecting Lateral Flow Assay

2021 ◽  
Author(s):  
David Cate ◽  
Helen Hsieh ◽  
Veronika Glukhova ◽  
Joshua D Bishop ◽  
H Gleda Hermansky ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Point Of Care ◽  
Lateral Flow ◽  
Screening Methods ◽  
Antibody Screening ◽  
Liquid Handling ◽  
Large Numbers ◽  
Accurate Performance ◽  
Further Development ◽  
Assay Conditions

<p></p><p>The global COVID-19 pandemic has created an urgent demand for large numbers of inexpensive, accurate, rapid, point-of-care diagnostic tests. Analyte-based assays are suitably inexpensive and can be rapidly mass-produced, but for sufficiently accurate performance they require highly optimized antibodies and assay conditions. We used an automated liquid handling system, customized to handle arrays of lateral flow immunoassay (LFA) tests in a high-throughput screen, to identify anti-nucleocapsid antibodies that will perform optimally in an LFA. We tested 1021 anti-nucleocapsid antibody pairs as LFA capture and detection reagents with the goal of highlighting pairs that have the greatest affinity for unique epitopes of the nucleocapsid protein of SARS-CoV-2 within the LFA format. In contrast to traditional antibody screening methods (e.g., ELISA, bio-layer interferometry), the method described here integrates real-time reaction kinetics with transport in, and immobilization directly onto, nitrocellulose. We have identified several candidate antibody pairs that are suitable for further development of an LFA for SARS-CoV-2.</p><p></p>

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