scholarly journals Vaccinia virus hemagglutinin. A novel member of the immunoglobulin superfamily.

1989 ◽  
Vol 170 (2) ◽  
pp. 571-576 ◽  
Author(s):  
D Y Jin ◽  
Z L Li ◽  
Q Jin ◽  
Y W Hao ◽  
Y D Hou
Keyword(s):  
Amino Acid ◽  
Vaccinia Virus ◽  
Functional Relationship ◽  
Immunoglobulin Superfamily ◽  
Hydrophobic Region ◽  
Acid Glycoprotein ◽  
Ig Superfamily ◽  
Infected Cells ◽  
Membrane Spanning ◽  
Human Cmv

Striking similarities between vaccinia virus hemagglutinin (VVHA) and proteins belonging to the Ig superfamily clearly indicate that VVHA, a 315-amino acid glycoprotein expressed on the surface of the infected cells, is a novel viral protein that can be added to the expanding list of the Ig superfamily. Its deduced amino acid sequence contains one Ig-like domain at the NH2 terminus, followed by two tandem repeating units and a hydrophobic region, suggestive of membrane spanning. The results offer an opportunity for the further study of the probable evolutionary and possible functional relationship between VVHA and other members of the Ig superfamily. Our observation, together with a recent finding that human CMV possibly encodes a protein similar to the MHC class I antigens (13), provides evidence supporting the fact that the viral capture of cellular Ig-related genes is more common than expected in vaccinia and other viruses, and that usage of an Ig-like domain as recognition signals might be extended from higher animals to animal viruses.

scholarly journals The Vaccinia Virus Superoxide Dismutase-Like Protein (A45R) Is a Virion Component That Is Nonessential for Virus Replication

2001 ◽  
Vol 75 (15) ◽  
pp. 7018-7029 ◽  
Author(s):  
Fernando Almazán ◽  
David C. Tscharke ◽  
Geoffrey L. Smith
Keyword(s):  
Superoxide Dismutase ◽  
Amino Acid ◽  
Vaccinia Virus ◽  
Virus Replication ◽  
Metal Ion ◽  
Deletion Mutant ◽  
Core Protein ◽  
Cowpox Virus ◽  
Sod Activity ◽  
Infected Cells

ABSTRACT A characterization of the A45R gene from vaccinia virus (VV) strain Western Reserve is presented. The open reading frame is predicted to encode a 125-amino-acid protein (M r, of 13,600) with 39% amino acid identity to copper-zinc superoxide dismutase (Cu-Zn SOD). Sequencing of the A45R gene from other orthopoxviruses, here and by others, showed that the protein is highly conserved in all viruses sequenced, including 16 strains of VV, 2 strains of cowpox virus, camelpox virus, and 4 strains of variola virus. In all cases the protein lacks key residues involved in metal ion binding that are important for the catalytic activity. The A45R protein was expressed inEscherichia coli, purified, and tested for SOD activity, but neither enzymatic nor inhibitory SOD activity was detected. Additionally, no virus-encoded SOD activity was detected in infected cells or purified virions. A monoclonal antibody raised against the A45R protein expressed in E. coli identified the A45R gene product as a 13.5-kDa protein that is expressed late during VV infection. Confocal microscopy of VV-infected cells indicated that the A45R protein accumulated predominantly in cytoplasmic viral factories. Electron microscopy and biochemical analyses showed that the A45R protein is incorporated into the virion core. A deletion mutant lacking the majority of the A45R gene and a revertant virus in which the deleted gene was restored were constructed and characterized. The growth properties of the deletion mutant virus were indistinguishable from those of wild-type and revertant viruses in all cell lines tested, including macrophages. Additionally, the virulence and pathogenicity of the three viruses were also comparable in murine and rabbit models of infection. A45R is unusual in being the first VV core protein described that affects neither virus replication nor virulence.

scholarly journals Membrane Organization of Bluetongue Virus Nonstructural Glycoprotein NS3

1998 ◽  
Vol 72 (4) ◽  
pp. 3362-3369 ◽  
Author(s):  
Om B. Bansal ◽  
Ann Stokes ◽  
Anju Bansal ◽  
David Bishop ◽  
Polly Roy
Keyword(s):  
Amino Acid ◽  
Cell Membrane ◽  
Vaccinia Virus ◽  
Bluetongue Virus ◽  
Expression System ◽  
Mature Protein ◽  
Nonstructural Proteins ◽  
Glycosylation Sites ◽  
Infected Cells ◽  
T7 Polymerase

ABSTRACT 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.

scholarly journals Cell surface anchorage and ligand-binding domains of the Saccharomyces cerevisiae cell adhesion protein alpha-agglutinin, a member of the immunoglobulin superfamily.

1993 ◽  
Vol 13 (4) ◽  
pp. 2554-2563 ◽  
Author(s):  
D Wojciechowicz ◽  
C F Lu ◽  
J Kurjan ◽  
P N Lipke
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Adhesion ◽  
Amino Acid ◽  
Cell Surface ◽  
Consensus Sequence ◽  
Binding Domain ◽  
Immunoglobulin Superfamily ◽  
Cell Contact ◽  
Amino Acid Residues ◽  
Adhesion Proteins

alpha-Agglutinin is a cell adhesion glycoprotein expressed on the cell wall of Saccharomyces cerevisiae alpha cells. Binding of alpha-agglutinin to its ligand a-agglutinin, expressed by a cells, mediates cell-cell contact during mating. Analysis of truncations of the 650-amino-acid alpha-agglutinin structural gene AG alpha 1 delineated functional domains of alpha-agglutinin. Removal of the C-terminal hydrophobic sequence allowed efficient secretion of the protein and loss of cell surface attachment. This cell surface anchorage domain was necessary for linkage to a glycosyl phosphatidylinositol anchor. A construct expressing the N-terminal 350 amino acid residues retained full a-agglutinin-binding activity, localizing the binding domain to the N-terminal portion of alpha-agglutinin. A 278-residue N-terminal peptide was inactive; therefore, the binding domain includes residues between 278 and 350. The segment of alpha-agglutinin between amino acid residues 217 and 308 showed significant structural and sequence similarity to a consensus sequence for immunoglobulin superfamily variable-type domains. The similarity of the alpha-agglutinin-binding domain to mammalian cell adhesion proteins suggests that this structure is a highly conserved feature of adhesion proteins in diverse eukaryotes.

scholarly journals A Combined Genetic-Proteomic Approach Identifies Residues within Dengue Virus NS4B Critical for Interaction with NS3 and Viral Replication

2015 ◽  
Vol 89 (14) ◽  
pp. 7170-7186 ◽  
Author(s):  
Laurent Chatel-Chaix ◽  
Wolfgang Fischl ◽  
Pietro Scaturro ◽  
Mirko Cortese ◽  
Stephanie Kallis ◽  
...  
Keyword(s):  
Amino Acid ◽  
Dengue Virus ◽  
Virus Replication ◽  
Strong Evidence ◽  
Drug Target ◽  
Nonstructural Protein ◽  
Rna Replication ◽  
Replication Cycle ◽  
Infected Cells ◽  
Ns3 Protease

ABSTRACTDengue virus (DENV) infection causes the most prevalent arthropod-borne viral disease worldwide. Approved vaccines are not available, and targets suitable for the development of antiviral drugs are lacking. One possible drug target is nonstructural protein 4B (NS4B), because it is absolutely required for virus replication; however, its exact role in the DENV replication cycle is largely unknown. With the aim of mapping NS4B determinants critical for DENV replication, we performed a reverse genetic screening of 33 NS4B mutants in the context of an infectious DENV genome. While the majority of these mutations were lethal, for several of them, we were able to select for second-site pseudoreversions, most often residing in NS4B and restoring replication competence. To identify all viral NS4B interaction partners, we engineered a fully viable DENV genome encoding an affinity-tagged NS4B. Mass spectrometry-based analysis of the NS4B complex isolated from infected cells identified the NS3 protease/helicase as a major interaction partner of NS4B. By combining the genetic complementation map of NS4B with a replication-independent expression system, we identified the NS4B cytosolic loop—more precisely, amino acid residue Q134—as a critical determinant for NS4B-NS3 interaction. An alanine substitution at this site completely abrogated the interaction and DENV RNA replication, and both were restored by pseudoreversions A69S and A137V. This strict correlation between the degree of NS4B-NS3 interaction and DENV replication provides strong evidence that this viral protein complex plays a pivotal role during the DENV replication cycle, hence representing a promising target for novel antiviral strategies.IMPORTANCEWith no approved therapy or vaccine against dengue virus infection, the viral nonstructural protein 4B (NS4B) represents a possible drug target, because it is indispensable for virus replication. However, little is known about its precise structure and function. Here, we established the first comprehensive genetic interaction map of NS4B, identifying amino acid residues that are essential for virus replication, as well as second-site mutations compensating for their defects. Additionally, we determined the NS4B viral interactome in infected cells and identified the NS3 protease/helicase as a major interaction partner of NS4B. We mapped residues in the cytosolic loop of NS4B as critical determinants for interaction with NS3, as well as RNA replication. The strong correlation between NS3-NS4B interaction and RNA replication provides strong evidence that this complex plays a pivotal role in the viral replication cycle, hence representing a promising antiviral drug target.

scholarly journals Vaccinia Virus F13L Protein with a Conserved Phospholipase Catalytic Motif Induces Colocalization of the B5R Envelope Glycoprotein in Post-Golgi Vesicles

2001 ◽  
Vol 75 (16) ◽  
pp. 7528-7542 ◽  
Author(s):  
Matloob Husain ◽  
Bernard Moss
Keyword(s):  
Vaccinia Virus ◽  
Fluorescent Protein ◽  
Plasmid Vector ◽  
Recombinant Vaccinia Virus ◽  
Open Reading Frames ◽  
Type I ◽  
Golgi Vesicles ◽  
Infected Cells ◽  
Catalytic Motif ◽  
Green Fluorescent

ABSTRACT The wrapping of intracellular mature vaccinia virions by modifiedtrans-Golgi or endosomal cisternae to form intracellular enveloped virions is dependent on at least two viral proteins encoded by the B5R and F13L open reading frames. B5R is a type I integral membrane glycoprotein, whereas F13L is an unglycosylated, palmitylated protein with a motif that is conserved in a superfamily of phospholipid-metabolizing enzymes. Microscopic visualization of the F13L protein was achieved by fusing it to the enhanced green fluorescent protein (GFP). F13L-GFP was functional when expressed by a recombinant vaccinia virus in which it replaced the wild-type F13L gene or by transfection of uninfected cells with a plasmid vector followed by infection with an F13L deletion mutant. In uninfected or infected cells, F13L-GFP was associated with Golgi cisternae and post-Golgi vesicles containing the LAMP 2 late endosomal-lysosomal marker. Association of F13L-GFP with vesicles was dependent on an intact phospholipase catalytic motif and sites of palmitylation. The B5R protein was also associated with LAMP2-containing vesicles when F13L-GFP was coexpressed, but was largely restricted to Golgi cisternae in the absence of F13L-GFP or when the F13L moiety was mutated. We suggest that the F13L protein, like its human phospholipase D homolog, regulates vesicle formation and that this process is involved in intracellular enveloped virion membrane formation.

Alterations of band 3 transport protein by cellular aging and disease: erythrocyte band 3 and glucose transporter share a functional relationship

1990 ◽  
Vol 68 (12) ◽  
pp. 1419-1427 ◽  
Author(s):  
Gieljan J. C. G. M. Bosman ◽  
Marguerite M. B. Kay
Keyword(s):  
Glucose Transport ◽  
Structural Changes ◽  
Glucose Transporter ◽  
Functional Relationship ◽  
Band 3 ◽  
Anion Transport ◽  
Cellular Aging ◽  
Abnormal Band ◽  
Membrane Spanning

Structural changes in human erythrocyte band 3 that affect anion transport are correlated with changes in glucose transport in situ. Breakdown of band 3, observed during normal erythrocyte aging in situ and in some diseases involving erythrocytes, is associated with an increase in Km and a decrease in Vmax of sulfate self-exchange, and with an increase in Km and Vmax of glucose efflux. Erythrocytes containing a high molecular weight form of band 3 exhibit an increase in Vmax of sulfate exchange and a decrease in Vmax of glucose efflux. Identical transport characteristics are observed in abnormal band-3-containing erythrocytes from individuals with familial amyotrophic chorea with acanthocytosis. A third band 3 alteration, fast-aging band 3, exhibits decreased Vmax of sulfate exchange and an increase in Km and decrease in Vmax of glucose efflux. Changes in band 3 structure that are the result of unstable hemoglobin or a deficiency in glucose-6-phosphate dehydrogenase and that do not affect anion transport have no effect on glucose transport characteristics. These data indicate the existence of a functional relationship between the membrane-spanning, anion-transport domain of band 3 and glucose transport in human erythrocytes. Antibodies to synthetic peptides reveal structural changes in membranes from the three inborn band 3 alterations and in band 3 itself in membranes from fast-aging band 3. Thus, immunological data suggests a structural relationship between anion and glucose transporters.Key words: red cell, anion transport, membrane proteins, aging, choreoacanthocytosis, anemia.

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