Nonspecific binding of immunoglobulins to coat proteins of certain plant viruses in immunoblots and indirect ELISA

We have reported that localization of TMV in tobacco cultivars with the N gene, is associated with a 23 K protein (IVR) that inhibited replication of several plant viruses. This protein was also found in induced resistant tissue of Nicotiana glutinosa x Nicotiana debneyi. During the present grant we found that TMV production is enhanced in protoplasts and plants of local lesion responding tobacco cultivars exposed to 35oC, parallel to an almost complete suppression of the production of IVR. We also found that IVR is associated with resistance mechanisms in pepper cultivars. We succeeded to clone the IVR gene. In the first attempt we isolated a clone - "101" which had a specific insert of 372 bp (the full length gene for the IVR protein of 23 kD should be around 700 bp). However, attempts to isolate the full length gene did not give clear cut results, and we decided not to continue with this clone. The amino acid sequence of the N-terminus of IVR was determined and an antiserum was prepared against a synthetic peptide representing amino acids residues 1-20 of IVR. Using this antiserum as well as our polyclonal antiserum to IVR a new clone NC-330 was isolated using lamba-ZAP library. This NC-330 clone has an insert of about 1 kB with an open reading frame of 596 bp. This clone had 86.6% homology with the first 15 amino acids of the N-terminal part of IVR and 61.6% homology with the first 23 amino acids of IVR. In the QIA expression system and western blotting of the expressed protein, a clear band of about 21 kD was obtained with IVR antiserum. This clone was used for transformation of Samsun tobacco plants and we have presently plantlets which were rooted on medium containing kanamycin. Hybridization with this clone was also obtained with RNA from induced resistant tissue of Samsun NN but not with RNA from healthy control tissue of Samsun NN, or infected or healthy tissue of Samsun. This further strengthens the previous data that the NC 330 clone codes for IVR. In the U.S. it was shown that IVR is induced in plants containing the N' gene when infected with mutants of TMV that elicit the HR. This is a defined system in which the elicitor is known to be due to permutations of the coat protein which can vary in elicitor strength. The objective was to understand how IVR synthesis is induced after recognition of elicitor coat protein in the signal transduction pathway that leads to HR. We developed systems to manipulate induction of IVR by modifying the elicitor and are using these elicitor molecules to isolate the corresponding plant receptor molecules. A "far-western" procedure was developed that found a protein from N' plants that specifically bind to elicitor coat proteins. This protein is being purified and sequenced. This objective has not been completed and is still in progress. We have reported that localization of TMV in tobacco cultivars with the N gene, is associated with a 23 K protein (IVR) that inhibited replication of several plant viruses. This protein was also found in induced resistant tissue of Nicotiana glutinosa x Nicotiana debneyi.

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Interaction of plant viruses and viral coat proteins with mixed model membranes

Biochemistry ◽

10.1021/bi00393a039 ◽

1987 ◽

Vol 26 (19) ◽

pp. 6217-6223 ◽

Cited By ~ 9

Author(s):

Klaas P. Datema ◽

Ruud B. Spruijt ◽

Benedictus J. M. Verduin ◽

Marcus A. Hemminga

Keyword(s):

Mixed Model ◽

Plant Viruses ◽

Model Membranes ◽

Coat Proteins ◽

Viral Coat ◽

Viral Coat Proteins

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A Unique Glycine-Rich Motif at the N-terminal Region of Bamboo mosaic virus Coat Protein Is Required for Symptom Expression

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-23-7-0903 ◽

2010 ◽

Vol 23 (7) ◽

pp. 903-914 ◽

Cited By ~ 30

Author(s):

Ping Lan ◽

Wen-Bin Yeh ◽

Chih-Wei Tsai ◽

Na-Sheng Lin

Keyword(s):

Mass Spectrometry ◽

Mosaic Virus ◽

Mutational Analysis ◽

Plant Viruses ◽

Site Directed Mutagenesis ◽

Mass Spectrometry Analysis ◽

Coat Proteins ◽

Long Distance ◽

Spectrometry Analysis ◽

Local Lesions

The coat proteins (CP) of many plant viruses are multifunctional proteins. We used N-terminal sequencing and mass spectrometry/mass spectrometry analysis to identify a truncated form of the Bamboo mosaic virus (BaMV) CP missing the N-terminal 35 amino acids (N35). The N35 region is unique in the potexviruses by its containing a glycine-rich motif (GRM) not present in databases but highly conserved among BaMV isolates. Results from site-directed mutagenesis and deletion mutational analysis showed that loss of this region converted necrotic local lesions to chlorotic local lesions on Chenopodium quinoa leaves. Furthermore, this region is required for successful development of mosaic symptoms on Nicotiana benthamiana leaves but is dispensable for BaMV replication and cell-to-cell and long-distance movement as well as virion assembly. This unique GRM-containing region of BaMV CP may be a symptom determinant in specific hosts.

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Structural Fingerprinting: Subgrouping of Comoviruses by Structural Studies of Red Clover Mottle Virus to 2.4-Å Resolution and Comparisons with Other Comoviruses

Journal of Virology ◽

10.1128/jvi.74.1.493-504.2000 ◽

2000 ◽

Vol 74 (1) ◽

pp. 493-504 ◽

Cited By ~ 23

Author(s):

Tianwei Lin ◽

Anthony J. Clark ◽

Zhongguo Chen ◽

Michael Shanks ◽

Jin-Bi Dai ◽

...

Keyword(s):

Mosaic Virus ◽

Red Clover ◽

Plant Viruses ◽

Small Subunit ◽

Mottle Virus ◽

Coat Proteins ◽

Structural Studies ◽

Bean Pod Mottle Virus ◽

X Ray ◽

N Terminus

ABSTRACT Red clover mottle virus (RCMV) is a member of the comoviruses, a group of picornavirus-like plant viruses. The X-ray structure of RCMV strain S has been determined and refined to 2.4 Å. The overall structure of RCMV is similar to that of two other comoviruses, Cowpea mosaic virus (CPMV) and Bean pod mottle virus (BPMV). The sequence of the coat proteins of RCMV strain O were modeled into the capsid structure of strain S without causing any distortion, confirming the close resemblance between the two strains. By comparing the RCMV structure with that of other comoviruses, a structural fingerprint at the N terminus of the small subunit was identified which allowed subgrouping of comoviruses into CPMV-like and BPMV-like viruses.

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Purification of plant viruses and virus coat proteins by high performance liquid chromatography

Journal of Virological Methods ◽

10.1016/0166-0934(90)90118-y ◽

1990 ◽

Vol 28 (3) ◽

pp. 245-256 ◽

Cited By ~ 6

Author(s):

Merete Albrechtsen ◽

Morten Heide

Keyword(s):

High Performance Liquid Chromatography ◽

Liquid Chromatography ◽

High Performance ◽

Plant Viruses ◽

Coat Proteins ◽

Virus Coat

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Measurement of anti-Aβ1–42 antibodies in intravenous immunoglobulin with indirect ELISA: The problem of nonspecific binding

Journal of Neuroscience Methods ◽

10.1016/j.jneumeth.2010.01.018 ◽

2010 ◽

Vol 187 (2) ◽

pp. 263-269 ◽

Cited By ~ 26

Author(s):

Andrea C. Klaver ◽

Lynnae M. Patrias ◽

Mary P. Coffey ◽

John M. Finke ◽

David A. Loeffler

Keyword(s):

Intravenous Immunoglobulin ◽

Indirect Elisa ◽

Nonspecific Binding

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The structure of a plant-specific partitivirus capsid reveals a unique coat protein domain architecture with an intrinsically disordered protrusion

Communications Biology ◽

10.1038/s42003-021-02687-w ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Matthew Byrne ◽

Aseem Kashyap ◽

Lygie Esquirol ◽

Neil Ranson ◽

Frank Sainsbury

Keyword(s):

Coat Protein ◽

Domain Architecture ◽

Plant Viruses ◽

Protein Domain ◽

Wild Plants ◽

Structural Basis ◽

Coat Proteins ◽

Intrinsically Disordered ◽

Binding Interface ◽

Protein Dimers

AbstractPersistent plant viruses may be the most common viruses in wild plants. A growing body of evidence for mutualism between such viruses and their hosts, suggests that they play an important role in ecology and agriculture. Here we present the capsid structure of a plant-specific partitivirus, Pepper cryptic virus 1, at 2.9 Å resolution by Cryo-EM. Structural features, including the T = 1 arrangement of 60 coat protein dimers, are shared with fungal partitiviruses and the picobirnavirus lineage of dsRNA viruses. However, the topology of the capsid is markedly different with protrusions emanating from, and partly comprising, the binding interface of coat protein dimers. We show that a disordered region at the apex of the protrusion is not required for capsid assembly and represents a hypervariable site unique to, and characteristic of, the plant-specific partitiviruses. These results suggest a structural basis for the acquisition of additional functions by partitivirus coat proteins that enables mutualistic relationships with diverse plant hosts.

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Structures of plant viruses from vibrational circular dichroism

Journal of General Virology ◽

10.1099/vir.0.81055-0 ◽

2005 ◽

Vol 86 (8) ◽

pp. 2371-2377 ◽

Cited By ~ 23

Author(s):

Ganesh Shanmugam ◽

Prasad L. Polavarapu ◽

Amy Kendall ◽

Gerald Stubbs

Keyword(s):

Circular Dichroism ◽

Coat Protein ◽

Mosaic Virus ◽

Structural Information ◽

Protein Structures ◽

Plant Viruses ◽

Vibrational Circular Dichroism ◽

Coat Proteins ◽

Helical Structures ◽

Circular Dichroïsm

Vibrational circular dichroism (VCD) spectra in the amide I and II regions have been measured for viruses for the first time. VCD spectra were recorded for films prepared from aqueous buffer solutions and also for solutions using D2O buffers at pH 8. Investigations of four filamentous plant viruses, Tobacco mosaic virus (TMV), Papaya mosaic virus, Narcissus mosaic virus (NMV) and Potato virus X (PVX), as well as a deletion mutant of PVX, are described in this paper. The film VCD spectra of the viruses clearly revealed helical structures in the virus coat proteins; the nucleic acid bases present in the single-stranded RNA could also be characterized. In contrast, the solution VCD spectra showed the characteristic VCD bands for α-helical structures in the coat proteins but not for RNA. Both sets of results clearly indicated that the coat protein conformations are dominated by helical structures, in agreement with earlier reports. VCD results also indicated that the coat protein structures in PVX and NMV are similar to each other and somewhat different from that of TMV. The present study demonstrates the feasibility of measuring VCD spectra for viruses and extracting structural information from these spectra.

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