Membrane organization of bluetongue virus nonstructural glycoprotein NS3.
The smallest RNA segment (S10) of bluetongue virus (an orbivirus, family Reoviridae) encodes two closely related nonstructural proteins, the 229-amino-acid (aa) NS3 and the 216-aa NS3A. The proteins are found in glycosylated and nonglycosylated forms in infected cells (X. Wu, H. Iwata, S.-Y. Chen, R. W. Compans and P. Roy J. Virol. 66:7104-7112, 1992). The NS3/NS3A proteins have two hydrophobic domains (aa 118 to 141 and 162 to 182) and two potential asparagine-linked glycosylation sites (aa 63 and 150), one of which is located between the hydrophobic domains. To determine whether these features were used in the mature protein forms, we generated a series of mutants of the S10 gene and expressed them by using the vaccinia virus T7 polymerase transient-expression system. Our data indicate that both hydrophobic domains of NS3 span the cell membrane and that only the site at aa 150 is responsible for N-linked glycosylation of the NS3 proteins.
Bansal
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Department of International Health, University of Alabama at Birmingham, 35294, USA.
Stokes
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Bansal
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Bishop
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Roy
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D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001820:Bluetongue virus / Q000235:genetics / Q000378:metabolism*; D050260:Carbohydrate Metabolism; D002522:Chlorocebus aethiops; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D006031:Glycosylation; D006056:Golgi Apparatus / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins
D011993:Recombinant Fusion Proteins; C075080:S10 protein, Bluetongue virus; D017361:Viral Nonstructural Proteins; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases