Interaction of murine macrophage-membrane proteins with components of the pathogenic fungus Histoplasma capsulatum.
The interaction of macrophage-membrane proteins and histoplasmin, a crude antigen of the pathogenic fungus Histoplasma capsulatum, was studied using murine peritoneal macrophages. Membrane proteins were purified via membrane attachment to polycationic beads and solubilized in Tris-HCl/SDS/DTT/glycerol for protein extraction; afterwards they were adsorbed or not with H. capsulatum yeast or lectin binding-enriched by affinity chromatography. Membrane proteins and histoplasmin interactions were detected by ELISA and immunoblotting assays using anti-H. capsulatum human or mouse serum and biotinylated goat anti-human or anti-mouse IgG/streptavidin-peroxidase system to reveal the interaction. Results indicate that macrophage-membrane proteins and histoplasmin components interact in a dose-dependent reaction, and adsorption of macrophage-membrane proteins by yeast cells induces a critical decrease in the interaction. Macrophage-membrane glycoproteins with terminal D-galactosyl residues, purified by chromatography with Abrus precatorius lectin, bound to histoplasmin; and two bands of 68kD and 180kD of transferred membrane protein samples interacted with histoplasmin components, as revealed by immunoblot assays. Specificity for beta-galactoside residues on the macrophage-membrane was confirmed by galactose inhibition of the interaction between macrophage-membrane proteins and histoplasmin components, in competitive ELISA using sugars, as well as by enzymatic cleavage of the galactoside residues.
Taylor
M L
ML
Department of Microbiología-Parasitología, Faculty of Medicine, UNAM, México, DF, Mexico.
Duarte-Escalante
E
E
Reyes-Montes
M R
MR
Elizondo
N
N
Maldonado
G
G
Zenteno
E
E
D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay; D006658:Histoplasma; D006659:Histoplasmin / Q000502:physiology; D006801:Humans; D015151:Immunoblotting; D008264:Macrophages / Q000276:immunology*; D008297:Male; D008565:Membrane Proteins / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C
D008565:Membrane Proteins; D006659:Histoplasmin