Monoclonal antibody affinity purification of a Leishmania membrane glycoprotein and its inhibition of leishmania-macrophage binding.
Specific monoclonal antibody coupled to Affi-Gel 10 was used to purify a major membrane glycoprotein of Leishmania mexicana amazonensis, one of a group of parasitic protozoa that specifically infect mammalian macrophages. Immobilized antigen was eluted at a 34% efficiency with buffers at either pH 2.5 or 11 or with MgCl2, but only the antigen eluted under basic conditions could be readsorbed to the immunobeads. Sephacryl S-300 gel filtration of the purified antigen gave a single peak of protein estimated to have a molecular mass of 400 kDa. However, NaDodSO4/polyacrylamide gel electrophoresis showed a single band of this protein with an apparent molecular mass of 63 kDa. The antigen is an N-linked glycoprotein, as indicated by its increase in electrophoretic mobility after treatment with endoglycosidase H and by its binding to lentil lectin-Sepharose, elutable with methyl alpha-D-mannoside and methyl alpha-D-glucoside. Purified antigen inhibits the binding of leishmania cells to macrophages by 50%, suggesting that it may play a role in the process of infection.
Chang
C S
CS
Chang
K P
KP
D000118:Acetylglucosaminidase / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000276:immunology*; D007158:Immunologic Techniques; D037102:Lectins; D007894:Leishmania mexicana / Q000276:immunology*; D008264:Macrophages / Q000276:immunology*; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D008970:Molecular Weight; D037121:Plant Lectins
D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D006023:Glycoproteins; D037102:Lectins; D037121:Plant Lectins; C033619:lentil lectin; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase