To establish its complete amino acid sequence we describe in this paper the results of sequence analysis on fragments derived from tetra-S-carboxymethyl-(CMA) and tetra-S-aminoethyl-(AEA) derivatives by chymotryptic, thermolytic and tryptic digestions. Parts of this work were published previously in Russian2-6).
ExperimentalMaterials and Reagents Actinoxanthin was isolated from the culture filtrate of Actinomyces globisporus 1131 via ammonium sulfate precipitation at 4-5°C followed by trichloroacetic acid precipitation of accompanied impurities , decolourizing by Dowex I x 2 (acetate form), chromatography using DEAE Sephadex A-50 (acetate form) and gel filtration on Sephadex G-75 as described earlier1). Analytical polyacrylamide gel electrophoresis was carried out using ORSTEIN-DAVIS System1). Homogeneity was shown by the following criteria: single band on polyacrylamide gel disc electrophoresis at pH 6.5, single symmetric peak by analytical column chromatography on Sephadex G-75, single NH,-terminal alanine by dansyl procedure , single C-terminal glycine by tritium-labeling procedure and amino acid analysis.Porcine trypsin was obtained from Novo (Denmark); chymotrypsin (crystallized) , thermolysin (3 x crystallized, 40 % Na-acetate, Ca-acetate), carboxypeptidases A and B (DFP-treated, in suspension) were obtained from Worthington Biochemical Corp., Freehold, NY, U.S.A.; leucine aminopeptidase was supplied by Serva Feinbiochemica Co., Heidelberg, FRG. All the enzymes were used without further purification.
The action of partly purified neuraminidase (NA) of influenza A virus, a mixture of detergent solubilized NA and haemagglutinin (HA) and of intact virions on gangliosides GTlb, GDla, GDlbr GM, was studied. The viral NA transformed GTlbmainly into GBlb with formation of only minor amounts of G,,. HA was found to inhibit the hydrolysis activity of viral NA. At the same time viral NA transformed Ga,,quantitatively into Ght, which was not hydrolyzed by the enzyme. These results suggest that the function of NA is to transfer the 'primary' receptor (such as GTlb) into the proper carbohydrate sequence (G,,,-like) which is proposed to serve as the minimal structure required for influenza virus reception.
Influenza virus receptor Viral neuraminidase Ganglioside
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