ORF 3 of Lactate Dehydrogenase-Elevating Virus Encodes a Soluble, Nonstructural, Highly Glycosylated, and Antigenic Protein

Open reading frame (ORF) 3 of the genome of lactate dehydrogenase-elevating virus (LDV), strain P, was cloned into the plasmid pcDNAI/Amp andin vitrotranscribed and translated. Translation of ORF 3 yielded a soluble protein of the expected size (about 21 kDa). When synthesized in the presence of end...

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Bibliographic Details
Published in:Virology (New York, N.Y.) N.Y.), 1997-01, Vol.227 (1), p.245-251
Main Authors: FAABERG, K.S., PLAGEMANN, P.G.W.
Format: Article
Language:English
Subjects:
Animals
Antibodies, Viral - immunology
Arterivirus Infections - immunology
Base Sequence
Cells, Cultured
Cloning, Molecular
COS Cells
DNA, Viral
Intracellular Membranes - virology
lactate dehydrogenase-elevating virus
Lactate dehydrogenase-elevating virus - genetics
Lactate dehydrogenase-elevating virus - immunology
Macrophages - virology
Mice
Microsomes - virology
Molecular Sequence Data
Open Reading Frames
Protein Sorting Signals - metabolism
Viral Nonstructural Proteins - genetics
Viral Nonstructural Proteins - immunology
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Summary:Open reading frame (ORF) 3 of the genome of lactate dehydrogenase-elevating virus (LDV), strain P, was cloned into the plasmid pcDNAI/Amp andin vitrotranscribed and translated. Translation of ORF 3 yielded a soluble protein of the expected size (about 21 kDa). When synthesized in the presence of endoplasmic reticulum (ER) membranes the resulting glycoprotein of about 36 kDa became associated with the membranes. However, disruption of the ER vesicles by incubation in carbonate buffer, pH 11.5, resulted in the release of the protein from the membranes. Hydrophobic moment analysis of the ORF 3 protein indicated the absence of any potential transmembrane segments, except for a N-terminal signal peptide, but no cleavage of the signal peptide was observed during membrane-associatedin vitrosynthesis. The ORF 3 protein elicited a strong antibody response in infected mice. The antibodies from infected mice as well as a monoclonal antibody specifically precipitated thein vitro-synthesized ORF 3 protein, but no protein from LDV virions. The overall results suggest that the ORF 3 protein is a nonstructural, highly glycosylated, and antigenic glycoprotein that is probably soluble and secreted or at most only weakly associated with membranes via the signal peptide.
ISSN:0042-6822
1096-0341
DOI:10.1006/viro.1996.8310