Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

In Vitro Expression and Production of Antibody Against Cymbidium mosaic virus Coat Protein.

ABSTRACT: Polyclonal rabbit antisera were produced using coat protein of Cymbidium mosaic virus (CymMV) Indian isolate expressed in E. coli as GST fusion. The expressed protein was purified by GST-fusion protein purification kit for use as an immunogen in rabbits. Antisera prepared in this manner reacted in double antibody sandwich enzyme-linked immunosorbent assay (DAS ELISA) with extract from CymMV-infected tissue. The results indicate that polyclonal antisera prepared from expressed CymMV coat proteins were useful for the detection of CymMV in an array of assays. The detection system developed is highly effective for detection of Indian strain of the virus in comparison to kits available in the international market.

SUBMITTER: Sherpa AR

PROVIDER: S-EPMC3550814 | biostudies-literature | 2012 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

In Vitro Expression and Production of Antibody Against Cymbidium mosaic virus Coat Protein.

Sherpa A R AR Hallan V V Zaidi A A AA

Indian journal of virology : an official organ of Indian Virological Society 20120427 1

Polyclonal rabbit antisera were produced using coat protein of Cymbidium mosaic virus (CymMV) Indian isolate expressed in E. coli as GST fusion. The expressed protein was purified by GST-fusion protein purification kit for use as an immunogen in rabbits. Antisera prepared in this manner reacted in double antibody sandwich enzyme-linked immunosorbent assay (DAS ELISA) with extract from CymMV-infected tissue. The results indicate that polyclonal antisera prepared from expressed CymMV coat proteins ...[more]

PMID: 23730003

Similar Datasets

Dual resistance of transgenic plants against Cymbidium mosaic virus and Odontoglossum ringspot virus.

Project description:Taxonomically distinct Cymbidium mosaic potexvirus (CymMV) and Odontoglossum ringspot tobamovirus (ORSV) are two of the most prevalent viruses worldwide; when co-infecting orchids, they cause synergistic symptoms. Because of the huge economic loss in quality and quantity in the orchid industry with virus-infected orchids, virus-resistant orchids are urgently needed. To date, no transgenic resistant lines against these two viruses have been reported. In this study, we generated transgenic Nicotiana benthamiana expressing various constructs of partial CymMV and ORSV genomes. Several transgenic lines grew normally and remained symptomless after mixed inoculation with CymMV and ORSV. The replication of CymMV and ORSV was approximately 70-90% lower in protoplasts of transgenic lines than wild-type (WT) plants. Of note, we detected extremely low or no viral RNA or capsid protein of CymMV and ORSV in systemic leaves of transgenic lines after co-infection. Grafting experiments further revealed that CymMV and ORSV trafficked extremely inefficiently from co-infected WT stocks to transgenic scions, presumably due to RNA-mediated interference. This study reports the first successful creation of dual resistant transgenic lines against CymMV and ORSV. Our studies shed light on the commercial development of transgenic orchid production to combat the global viral threat.

| S-EPMC6629631 | biostudies-literature

Alfalfa mosaic virus coat protein bridges RNA and RNA-dependent RNA polymerase in vitro.

Project description:Alfalfa mosaic virus (AMV) RNA replication requires the viral coat protein (CP). AMV CP is an integral component of the viral replicase; moreover, it binds to the viral RNA 3'-termini and induces the formation of multiple new base pairs that organize the RNA conformation. The results described here suggest that AMV coat protein binding defines template selection by organizing the 3'-terminal RNA conformation and by positioning the RNA-dependent RNA polymerase (RdRp) at the initiation site for minus strand synthesis. RNA-protein interactions were analyzed by using a modified Northwestern blotting protocol that included both viral coat protein and labeled RNA in the probe solution ("far-Northwestern blotting"). We observed that labeled RNA alone bound the replicase proteins poorly; however, complex formation was enhanced significantly in the presence of AMV CP. The RNA-replicase bridging function of the AMV CP may represent a mechanism for accurate de novo initiation in the absence of canonical 3' transfer RNA signals.

| S-EPMC2583179 | biostudies-literature

Characterization of Alternanthera mosaic virus and its Coat Protein.

Project description:A new isolate of Alternantheramosaic virus (AltMV-MU) was purified from Portulaca grandiflora plants. It has been shown that the AltMV-MU coat protein (CP) can be efficiently reassembled in vitro under different conditions into helical RNA-free virus-like particles (VLPs) antigenically related to native virus. The AltMV-MU and VLPs were examined by atomic force and transmission electron microscopies. The encapsidated AltMV-MU RNA is nontranslatable in vitro. However, it can be translationally activated by CP phosphorylation or by binding to the TGB1protein from the virus-coded movement triple gene block.

| S-EPMC3245411 | biostudies-literature

Nuclear targeting of the cauliflower mosaic virus coat protein.

Project description:The entry of the viral genomic DNA of cauliflower mosaic virus into the nucleus is a critical step of viral infection. We have shown by transient expression in plant protoplasts that the viral coat protein (CP), which is processed from the product of open reading frame IV, contains an N-terminal nuclear localization signal (NLS). The NLS is exposed on the surface of the virion and is thus available for interaction with a putative NLS receptor. Phosphorylation of the matured CP did not influence the nuclear localization of the protein but improved protein stability. Mutation of the NLS completely abolished viral infectivity, thus indicating its importance in the virus life cycle. The NLS seems to be regulated by the N terminus of the precapsid, which inhibits its nuclear targeting. This regulation could be important in allowing virus assembly in the cytoplasm.

| S-EPMC103861 | biostudies-other

Cymbidium mosaic potexvirus isolate-dependent host movement systems reveal two movement control determinants and the coat protein is the dominant.

Project description:Little is known about how plant viruses of a single species exhibit different movement behavior in different host species. Two Cymbidium mosaic potexvirus (CymMV) isolates, M1 and M2, were studied. Both can infect Phalaenopsis orchids, but only M1 can systemically infect Nicotiana benthamiana plants. Protoplast inoculation and whole-mount in situ hybridization revealed that both isolates can replicate in N. benthamiana; however, M2 was restricted to the initially infected cells. Genome shuffling between M1 and M2 revealed that two control modes are involved in CymMV host dependent movement. The M1 coat protein (CP) plays a dominant role in controlling CymMV movement between cells, because all chimeric CymMV viruses containing the M1 CP systemically infected N. benthamiana plants. Without the M1 CP, one chimeric virus containing the combination of the M1 triple gene block proteins (TGBps), the M2 5' RNA (1-4333), and the M2 CP effectively moved in N. benthamiana plants. Further complementation analysis revealed that M1 TGBp1 and TGBp3 are co-required to complement the movement of the chimeric viruses in N. benthamiana. The amino acids within the CP, TGBp1 and TGBp3 which are required or important for CymMV M2 movement in N. benthamiana plants were mapped. The required amino acids within the CP map to the predicted RNA binding domain. RNA-protein binding assays revealed that M1 CP has higher RNA binding affinity than does M2 CP. Yeast two-hybrid assays to detect all possible interactions of M1 TGBps and CP, and only TGBp1 and CP self-interactions were observed.

| S-EPMC7103407 | biostudies-literature

Cofolding organizes alfalfa mosaic virus RNA and coat protein for replication.

Project description:Alfalfa mosaic virus genomic RNAs are infectious only when the viral coat protein binds to the RNA 3' termini. The crystal structure of an alfalfa mosaic virus RNA-peptide complex reveals that conserved AUGC repeats and Pro-Thr-x-Arg-Ser-x-x-Tyr coat protein amino acids cofold upon interacting. Alternating AUGC residues have opposite orientation, and they base pair in different adjacent duplexes. Localized RNA backbone reversals stabilized by arginine-guanine interactions place the adenosines and guanines in reverse order in the duplex. The results suggest that a uniform, organized 3' conformation, similar to that found on viral RNAs with transfer RNA-like ends, may be essential for replication.

| S-EPMC1500904 | biostudies-literature

Phylodynamics and Coat Protein Analysis of Babaco Mosaic Virus in Ecuador.

Project description:Babaco is a fast-growing herbaceous shrub with great commercial potential because of the organoleptic properties of its fruit. Babaco mosaic virus (BabMV) is a potexvirus in the family Alphaflexiviridae affecting babaco in all the provinces that produce this crop in Ecuador. BabMV was recently described but it has been affecting babaco for decades and, since many potexviruses are serologically indistinguishable, it may have been previously misidentified as papaya mosaic virus. Based on the coat protein (CP) gene, we aimed to study the distribution and epidemiological patterns of BabMV in babaco and chamburo over the years and to model its three-dimensional structure. Sequences of the CP were obtained from thirty-six isolates from plants collected in the main babaco-producing provinces of Ecuador between 2016 and 2021. The evolution rate of BabMV was estimated at 1.21 × 10-3 nucleotide substitutions site-1 year-1 and a time of origin of the most recent common ancestor around 1958.80. From molecular dynamics simulations, compared to other proteins of BabMV-RDRP, TGB1, and Alkb domain-the CP exhibited a higher flexibility with the C and N terminals as the most flexible regions. The reconstructed viral distribution provides dispersion patterns which have implications for control approaches of BabMV.

| S-EPMC9268947 | biostudies-literature

Spatial determinants of the alfalfa mosaic virus coat protein binding site.

Project description:The biological functions of RNA-protein complexes are, for the most part, poorly defined. Here, we describe experiments that are aimed at understanding the functional significance of alfalfa mosaic virus RNA-coat protein binding, an interaction that parallels the initiation of viral RNA replication. Peptides representing the RNA-binding domain of the viral coat protein are biologically active in initiating replication and bind to a 39-nt 3'-terminal RNA with a stoichiometry of two peptides: 1 RNA. To begin to understand how RNA-peptide interactions induce RNA conformational changes and initiate replication, the AMV RNA fragment was experimentally manipulated by increasing the interhelical spacing, by interrupting the apparent nucleotide symmetry, and by extending the binding site. In general, both asymmetric and symmetric insertions between two proposed hairpins diminished binding, whereas 5' and 3' extensions had minimal effects. Exchanging the positions of the binding site hairpins resulted in only a moderate decrease in peptide binding affinity without changing the hydroxyl radical footprint protection pattern. To assess biological relevance in viral RNA replication, the nucleotide changes were transferred into infectious genomic RNA clones. RNA mutations that disrupted coat protein binding also prevented viral RNA replication without diminishing coat protein mRNA (RNA 4) translation. These results, coupled with the highly conserved nature of the AUGC865-868 sequence, suggest that the distance separating the two proposed hairpins is a critical binding determinant. The data may indicate that the 5' and 3' hairpins interact with one of the bound peptides to nucleate the observed RNA conformational changes.

| S-EPMC1370517 | biostudies-literature

Alcohol-perturbed self-assembly of the tobacco mosaic virus coat protein.

Project description:The self-assembly of the tobacco mosaic virus coat protein is significantly altered in alcohol-water mixtures. Alcohol cosolvents stabilize the disk aggregate and prevent the formation of helical rods at low pH. A high alcohol content favours stacked disk assemblies and large rafts, while a low alcohol concentration favours individual disks and short stacks. These effects appear to be caused by the hydrophobicity of the alcohol additive, with isopropyl alcohol having the strongest effect and methanol the weakest. We discuss several effects that may contribute to preventing the protein-protein interactions between disks that are necessary to form helical rods.

| S-EPMC8978915 | biostudies-literature

Degenerate in vitro genetic selection reveals mutations that diminish alfalfa mosaic virus RNA replication without affecting coat protein binding.

Project description:The alfalfa mosaic virus (AMV) RNAs are infectious only in the presence of the viral coat protein; however, the mechanisms describing coat protein's role during replication are disputed. We reasoned that mechanistic details might be revealed by identifying RNA mutations in the 3'-terminal coat protein binding domain that increased or decreased RNA replication without affecting coat protein binding. Degenerate (doped) in vitro genetic selection, based on a pool of randomized 39-mers, was used to select 30 variant RNAs that bound coat protein with high affinity. AUGC sequences that are conserved among AMV and ilarvirus RNAs were among the invariant nucleotides in the selected RNAs. Five representative clones were analyzed in functional assays, revealing diminished viral RNA expression resulting from apparent defects in replication and/or translation. These data identify a set of mutations, including G-U wobble pairs and nucleotide mismatches in the 5' hairpin, which affect viral RNA functions without significant impact on coat protein binding. Because the mutations associated with diminished function were scattered over the 3'-terminal nucleotides, we considered the possibility that RNA conformational changes rather than disruption of a precise motif might limit activity. Native polyacrylamide gel electrophoresis experiments showed that the 3' RNA conformation was indeed altered by nucleotide substitutions. One interpretation of the data is that coat protein binding to the AUGC sequences determines the orientation of the 3' hairpins relative to one another, while local structural features within these hairpins are also critical determinants of functional activity.

| S-EPMC446135 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data