Characterization of receptors for encephalomyocarditis virus on cells

Baldeh, Mansajang P. (1987) Characterization of receptors for encephalomyocarditis virus on cells. Masters thesis, Memorial University of Newfoundland.

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Abstract

A great deal more is known about viruses than about the receptor structures that allow entry of viruses into cells. The work reported in this thesis was aimed at finding out more about the structure of a receptor for the mouse encephalomyocarditis (EMC) virus that is found on some mouse cells and on human erythroid cells. Since previous reports clarified that the receptor for EMC virus on the human crythrocyte is glycophorin A (Allaway et al., 1986), a virus-Sepharose column was used to study the binding of glycophorin to the column as well as to study binding between the virus and its surface receptors on human erythrocytes and Krebs II ascites tumour cells. Glycophorin was retained on the virus-Sepharose column and was eluted with 0.02 M sodium phosphate buffer pII 8.0 containing 0.02 to 0.05 M NaCI. This result indicates that weak ionie interactions are involved between the virus and glycophorin. -- Radio-labelled molecules from red cell membranes solubilized in detergent were found to bind to EMC virus-Sepharose and analysis of eluates by electrophoresis showed that the bound material contained both glycophorins A and B. When glycophorins A and B isolated by gel filtration on Biogel columns were separately applied to the EMC virus-Sepharose column, both sialoglycoproteins (glycophorius A and B) were retained on the virus column. Contrary to previous reports this result suggests that both glycophorins A and B may serve as receptors for EMC virus. -- Radio-labelled molecules from red cell membranes solubilized in detergent were found to bind to EMC virus-Sepharose and analysis of eluates by electrophoresis showed that the bound material contained both glycophorins A and B. When glycophorins A and B isolated by gel filtration on Biogel columns were separately applied to the EMC virus-Sepharose column, both sialoglycoproteins (glycophorins A and B) were retained on the virus column. Contrary to previous reports this result suggests that both glycophorins A and B may serve as receptors for EMC virus. -- Radio-labelled Krebs cell membranes were solubilized in detergent and applied to the virus-Sepharose column. The column eluates analysed by SDS-PAGE showed a peak at the origin of the gel and this material was sensitive to trypsin treatment but not treatment with neuraminidase which releases sialic acids. This result suggests that the receptor for EMC virus on the Krebs cell may be a non sialylated glycoprotein. -- In a second approach, to characterize the receptor for EMC virus on cells, anti-idiotypic antibodies were used. Antibodies to the EMC virus were raised in New Zealand White rabbits; these antibodies were purified by affinity chromatography and then injected into the same breed of rabbits to produce anti-idiotypic antibodies. Such antibodies should mimic the binding properties of the virus itself. The anti-idiotypic antibodies were found to: -- 1. bind to glycophorin immobilised on nitrocellulose. -- 2. immunoprecipitate blycophorin from solubilized radiolabelled red cell membranes. -- 3. agglutinate both human and sheep erythrocytes but not bovine nor rabbit red blood cells. -- These results suggest that anti-idiotypic antibodies against EMC virus have been successfully made and these antibodies bind to the receptor for EMC virus on human erythorocytes.

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