Enzymatic Iodination of Sindbis Virus Proteins

Sefton, Bartholomew M.; Wickus, Gary G.; Burge, Boyce W.

doi:10.1128/jvi.11.5.730-735.1973

Cited by 98 publications

(61 citation statements)

References 21 publications

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“…Phinney et al [55] also found that E2 protected E1 from trypsinisation and iodination at neutral pH although E1 became highly trypsin sensitive after exposure of the virus to acid pH. This is in agreement with earlier studies since iodination was known to detect more tyrosine on E2 than on E1 [72]. Biotinylation studies of exposed lysines on the surface of infectious Sindbis virus identified several sites on E2 (K70, 76,97,131,149,202,235) confirming the location of aa 202 in the exposed distal tip cluster [73].…”

Section: Figure 2 (A) a Non-reducing Page Of E1 And E2 Extracted Fromsupporting

confidence: 75%

Sindbis virus as a model for studies of conformational changes in a metastable virus and the role of conformational changes in in vitro antibody neutralisation

Hernandez

Paredes

2009

Reviews in Medical Virology

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Sindbis virus has long been the prototypical alphavirus used in the study of assembly of icosahedral viruses which contain membranes. Much is known about the assembly pathway and molecular architecture of this virus, and models of mechanisms of infection and release of the viral RNA have been proposed. This review will focus on the structural aspects of in vitro antibody neutralisation of a metastable alphavirus, Sindbis virus, which results from antibody induced conformational changes. For the alphaviruses, structures of in vitro antibody induced conformational neutralisation for Sindbis virus, and receptor occlusion for Sindbis and Ross River virus (RRV) have been reported. A model is presented which could extend the mechanism of antibody induced conformational neutralisation to any metastable virus structure.

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Section: Figure 2 (A) a Non-reducing Page Of E1 And E2 Extracted Fromsupporting

confidence: 75%

Sindbis virus as a model for studies of conformational changes in a metastable virus and the role of conformational changes in in vitro antibody neutralisation

Hernandez

Paredes

2009

Reviews in Medical Virology

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“…The lactoperoxidase catalyzed iodination reaction (Hubbard and Cohn, 1972;Sefton et al, 1973) was carried out directly in the culture medium to preserve the natural conditions and to avoid any alteration induced by washing procedures.…”

Section: Surjkce Ioditiation Oj' Cellsmentioning

confidence: 99%

Fibroblast surface antigen (SF): THE external glycoprotein lost in proteolytic stimulation and malignant transformation

Keski‐Oja

Vaheri

Ruoslahti

1976

Intl Journal of Cancer

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It was previously shown that the fibroblast surface antigen (SF antigen, SFA) is composed of polypeptides of high molecular weight 210,000 (SF210) and 145,000 (SF145) and that both of these decrease in quantity after transformation of the fibroblasts by Rous sarcoma virus (RSV). The present experiments show that SF210 is a glycoprotein. It is accessible to surface labelling by lactoperoxidase catalyzed iodination. The SF210 molecule is highly susceptible to trypsin on cell surface. Anti-SFA antibodies specifically precipitated the surface labelled polypeptide. The lactoperoxidase iodinated SF210 polypeptide was greatly reduced in cells transformed by RSV. It is concluded from these studies that the large external transformation sensitive (LETS) protein detected by other workers is the same molecule as SF210. Part of the label of surface iodinated fibroblasts did not enter the polyacrylamide gels. This high molecular weight material is also susceptible to trypsin treatment and decreases in quantity after transformation by RSV. The data suggest that it may be antigenically related to SF protein. Treatment of surface of 35S-methionine-labelled cultures with trypsin in concentrations able to initiate proliferation of density-inhibited cells rapidly released SF210 from fibroblast surface. A single high molecular weight polypeptide (mol. wt about 200,000, SF200) was detected in the culture medium. SF210 may thus be a major target molecule of trypsin action. Treatment of cultures with insulin that also stimulated the fibroblasts to initiate proliferation did not result in any detectable alteration in the external glycoprotein SF210. It is concluded that although release of SF210 may be a sufficient trigger to stimulate proliferation in stationary cells, this molecule appears not to be directly involved in initiation of fibroblast proliferation from the G1 (or G0) phase of the cell cycle.

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“…0.2 ml packed cells was resuspended in 1.8 ml buffer A. Iodination with 50 taCi [l*q]NaI was performed as detailed by Sefton et al (29) for Sindbis virus-infected cells, except that iodination was performed in buffer A. The cells were recovered by centrifugation, and washed four times by centrifugation in buffer A.…”

Section: Iodination Of Cellsmentioning

confidence: 99%

Membrane proteins synthesized by rabbit reticulocytes.

Lodish¹,

Small²

1975

The Journal of Cell Biology

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Intact rabbit reticulocyte cells synthesize two predominant species of polypeptides which are components of the cell plasma membrane. Previous work (Lodish, H. F. 1973. Proc. Natl. Acad. Sci. U. S. A. 70:1526-1530 showed that these proteins were synthesized by polyribosomes not attached to membranes. We show here that both polypeptides are confined to the cytoplasmic surface of the cell membrane. These studies utilized iodination of whole cells and of membranes with lactoperoxidase, and digestion of whole cells and membranes with chymotrypsin. One of these proteins is synthesized as a precursor, and about 20-40 amino acids are removed after it is incorporated into the membrane. We discuss the probable sites of synthesis of these and other classes of membrane proteins.Although a great deal is known of the protein composition of mammalian erythrocyte membranes, and of the asymmetric distribution of these proteins within the membrane, little is known about the biosynthesis of erythrocyte membrane proteins. The literature concerning the proteins of erythrocyte membranes has been reviewed several times (1-4). As resolved by sodium dodecyl sulfate (SDS) gel electrophoresis, all mammalian erythrocyte membranes contain about seven to nine principal protein components, although the detailed pattern of membrane proteins shows some species differences (1-11). The major polypeptides common to all mammalian erythrocytes include two polypeptides of molecular weight greater than 200,000 (7) and one of 43,000 (the three spectrin polypeptides) (1, 3, 4, 12), a protein of molecular weight 90,000, and a sialoylglycoprotein which in the human contains the MN, A, and B blood group antigens (11). The latter two species are the only membrane proteins found on the cell surface; recent work suggests that they penetrate to the interior surface of the membrane (references 6, 13-20, reviewed in reference 4). Most membrane proteins are confined to the cytoplasmic surface of the membrane; glyceraldehyde-3-phosphate dehydrogenase, for instance, is found only on the inner surface of the human erythrocyte membrane, and the isolated enzyme forms stable interactions with high affinity for a limited number of sites on the cytoplasmic face of the membrane (21).Rabbit reticulocytes contain, with possibly one or two exceptions (22), the same proteins in their membranes as do rabbit erythrocytes. We recently showed that rabbit reticulocytes synthesize only two major and two to three minor membrane proteins; most of the membrane proteins are no longer being synthesized in significant amounts at this stage (23). We also showed that these two proteins as well as all other proteins made by reticulocytes are synthesized on polyribosomes which are not attached to membranes (23,24). A small fraction of reticulocyte polysomes are apparently bound to the cell membrane. Woodward et

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Enzymatic Iodination of Sindbis Virus Proteins

Cited by 98 publications

References 21 publications

Sindbis virus as a model for studies of conformational changes in a metastable virus and the role of conformational changes in in vitro antibody neutralisation

Sindbis virus as a model for studies of conformational changes in a metastable virus and the role of conformational changes in in vitro antibody neutralisation

Fibroblast surface antigen (SF): THE external glycoprotein lost in proteolytic stimulation and malignant transformation

Membrane proteins synthesized by rabbit reticulocytes.

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