Radioiodination and identification of externally disposed membrane components of Leishmania tropica.
Two methods, one involving a lactoperoxidase-glucose oxidase coupled reaction and the other employing the insoluble catalyst 1,3,4,6-tetrachloro-3 alpha, 6-alpha-diphenyl-glycoluril (Iodo-Gen), were used to label the surface membrane of promastigotes of Leishmania tropica major. Both methods labelled approximately 20 proteins or glycoproteins (apparent size range 10 to 110 kDa) in a qualitatively similar manner, however the lactoperoxidase method labelled one additional constituent (260 kDa). By omission of both enzymes, or of Iodo-Gen; by comparison of radioactivity incorporated by particulate and soluble cell fractions; and through the action of proteases on live, labelled promastigotes, the surface-labelling specificity of both procedures was confirmed. Immunoprecipitation of Triton X-100 extracts of labelled cells with rabbit antisera revealed a minimum of twelve (seven major) protein antigens in the homologous system and different but cross-reactive protein species from two other isolates of L. tropica. Lectin precipitation of radiolabelled surface components was possible with concanavalin A (but not with other lectins tested) identifying a minimum of 12 glycoproteins. Two of these glycoproteins (210 and 88 kDa) were not recognized by rabbit antiserum.
D000818:Animals; D001345:Autoradiography; D002462:Cell Membrane / Q000648:ultrastructure; D011232:Chemical Precipitation; D007120:Immunochemistry; D007457:Iodine Radioisotopes; D007784:Lactoperoxidase; D037102:Lectins; D007891:Leishmania / Q000648:ultrastructure*; D010447:Peptide Hydrolases
D007457:Iodine Radioisotopes; D037102:Lectins; D007784:Lactoperoxidase; D010447:Peptide Hydrolases