The possibility that the haemagglutinin of encephalomyocarditis (EMC) virus might be a glycoprotein was investigated by looking for sugars and amino sugars in purified EMC virus particles grown in Krebs ascitestumour cells in vitro in the presence of isotopically labelled glucosamine, acetyl glucosamine, galactosamine, mannosamine or fucose. With either isotopically labelled glucosamine or galactosamine the virus became significantly radioactive, whereas radioactivity was low in virusgrown in the presence of mannosamine, or fucose, possibly because the latter was poorly taken up by the cells; acetyl glucosamine was completely excluded by Krebs cells, so its ability to be incorporated into virus could not be tested.
Radioactivity derived from glucosamine remained firmly attached to the purified virus through both rate-zonal and equilibrium density centrifugation and chromatography on calcium phosphate. This radioactivity did not appear due to simple adsorption of either glucosamine or (after synthesis) acetyl glucosamine, or to contamination of the preparation with infected host cell membranes. Estimates of the number of glucosamine molecules incorporated into the virus had values which varied with the position of the isotope in the glucosamine added, suggesting that it was not incorporated unchanged.
Polyacrylamide gel electrophoresis of disrupted whole virus showed radio-activity derived from glucosamine to be present in all virus polypeptides, indicating that glucosamine was perhaps metabolized, amongst other things, to amino acids. Disruption of the virus with phenol revealed that 70 % of the radioactivity was in the virus RNA, mostly in ribose, the remaining 30% being in virus protein, presumably as amino acids. The total amount of radioactivity that could be in the form of glucosamine did not allow for more than 0.6 glucosamine residues per virus particle. From this it was concluded that glucosamine is not a virus constituent per se, that the EMC virus particle does not contain glycoprotein and, consequently, that the virus haemagglutinin is not a glycoprotein.
BellettA. J. D.,
BurnessA. T. H.1963; Intracellular sites of synthesis of encephalomyocarditis virus components in Krebs-2 ascites tumour cells. Journal of General Microbiology 30:131–140
BrunerR.,
VinogradJ.1965; The evaluation of standard sedimentation coefficients of sodium RNA and sodium DNA from sedimentation velocity data in concentrated NaCl and CsCl solutions. Bio-chimica et biophysica acta 108:18–29
BurnessA. T. H.1967; Separation of plaque-type variants of encephalomyocarditis virus by chromatography on calcium phosphate. Journal of Virology 1:308
BurnessA. T. H.1969a; Purification and separation of encephalmoyocarditis virus variants by chromatography on calcium phosphate.. In Fundamental Techniques in Virology Edited by
HabelK.,
SalzmanN.
New York and London: Academic Press;
BurnessA. T. H.,
ClothierF. W.1970; Particle weight and other biophysical properties of encephalomyocarditis virus. Journal of General Virology 6:381–393
CompansR. W.,
KlenkH. D.,
CaliguiriL. A.,
ChoppinP. W.1970; Influenza virus proteins. I. Analysis of polypeptides of the virion and identification of spike glycoproteins. Virology 42:880–889
DavidsonE. A.1966; Analysis of sugars found in mucopolysaccharides. In Methods in Enzymology8 Edited by
ColowickS. P.,
KaplanN. O.,
MenfeldE. F.,
GinsbergV.
New York and London: Academic Press;
HalperenS.,
StoneH. O.,
KorantB. D.1970Isolation of glucosamine from the capsids of picornavirus. Abstract V18Bacteriological Proceedings, 70th Meeting of American Society for Microbiology Boston, Mass., U.S.A:
HaslamE. A.,
HampsonA. W.,
EganJ. A.,
WhiteD. O.1970a; The polypeptides of influenza virus. II. Interpretation of polyacrylamide gel electrophoresis patterns. Virology 42:555–565
HaslamE. A.,
hampsonA. W.,
RadiskevisI.,
WhiteD. O.1970b; The polypeptides of influenza virus. III. Identification of the hemagglutinin, neuraminidase and nucleocapsid proteins. Virology 42:566–575
HoskinsJ. M.,
SandersF. K.1957; Propagation of mouse encephalomyocarditis virus in ascitestumour cells maintained in vitro. British Journal of Experimental Pathology 38:268–272
International Critical Tables of Numerical Data, Physics, Chemistry, Technology; 1927; 2337 Edited by
WashbumE. W.
et al New York and London: McGraw-Hill;
KlenkH. D.,
CaliguiriL. A.,
ChoppinP. W.1970; The proteins of the parainfluenza virus SV5. II. The carbohydrate content and glycoproteins of the virion. Virology 42:473–481
MarkhamR.,
SmithJ. D.1952; The structure of ribonucleic acids. I. Cyclic nucleotides produced by ribonuclease and by alkaline hydrolysis. Biochemical Journal 52:552–557
RueckertR. R.1965; Studies on the structure of viruses of the Columbia SK Group. II. The protein sub-units of ME-virus and other members of the Columbia SK Group. Virology 26:345–358
SchwerdtC. E.1959 Quantitative relationships between virus particles and their functional activity.. In The Viruses1329 Edited by
BurnetF. M.,
StanleyW. M.
New York and London: Academic Press;
TillotsonJ. R.,
LernerA. M.1966; Effect of periodate oxidation on haemagglutinating and antibodyproducing capacities of certain enteroviruses and reoviruses. Proceedings of the National Academy of Sciences of the United States of America 56:1143–1150
WheatR. W.1966 Analysis of hexosamines in bacterial polysaccharides by chromatographic procedures. In Methods in Enzymology8 Edited by
ColowickS. P.,
KaplanN. O.,
MenfeldE. F.,
GinsbergV.
New York and London: Academic Press;
WyattG. R.1955 Separation of nucleic acid components by chromatography on filter paper.. In The Nucleic Acids Edited by
ChargaffE.,
DavidsonJ. N.
New York and London: Academic Press;