Electron microscopic identification of exposed plasmin epitopes in alpha 2-macroglobulin-plasmin complex using monoclonal antibody-colloidal gold adducts.
Monoclonal antibodies 10-F-1, directed against the K4 region of plasminogen, and 10-V-1, directed against the K1-3 region of plasminogen, were adducted to colloidal gold. These antibody-gold adducts bound specifically to alpha 2-macroglobulin (alpha 2M)-plasmin. Greater than 90% of the apparent binding was eliminated when alpha 2M-methylamine was substituted for the alpha 2M-plasmin. The plasmin epitope recognized by 10-F-1 was identified at the extreme pole of the alpha 2M-plasmin complex, suggesting that plasmin protrudes from the end of the hollow cylinder formed by alpha 2M. The complexes formed between alpha 2M-plasmin and 10-V-1 were indistinguishable from those formed with 10-F-1. This suggests that exposure of plasmin surface structure in alpha 2M-plasmin, while substantial, may be limited to the single region of the inhibitor. Evidence for ternary complexes containing one alpha 2M and two plasmin molecules was obtained in the form of antibody-gold bound at both poles of alpha 2M-plasmin. The fraction of alpha 2M-plasmin that associated with more than one antibody was small. The data presented here are considered in relation to current models of alpha 2M structure and function.
Gonias
S L
SL
Department of Pathology, University of Virginia Medical Center, Charlottesville 22908.
Allietta
M M
MM
Pizzo
S V
SV
Castellino
F J
FJ
Tillack
T W
TW
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000939:Epitopes / Q000032:analysis*; D005341:Fibrinolysin / Q000276:immunology / Q000378:metabolism*; D006046:Gold; D006801:Humans; D008854:Microscopy, Electron; D011485:Protein Binding; D011487:Protein Conformation; D000511:alpha-Macroglobulins / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000939:Epitopes; D000511:alpha-Macroglobulins; D006046:Gold; D005341:Fibrinolysin