IMMUNOELECTRON MICROSCOPY STUDIES WITH A MONOCLONAL-ANTIBODY DIRECTED AGAINST A RECEPTOR RECOGNITION SITE EPITOPE IN HUMAN ALPHA-2-MACROGLOBULIN

α2-Macroglobulin (α2M) is a plasma proteinase inhibitor that binds up to 2 mole of proteinase per mole of inhibitor. Proteinase binding or reaction with small primary amines causes a major conformational change in α2M. As a result of this conformational change, a new epitope recognized by monoclonal antibody 7H11D6 is exposed. The association of α2M-proteinase or α2M-methylamine with α2M cellular receptors is prevented by 7H11D6. In this investigation, the binding of 7H11D6 to α2M was studied by electron microscopy. 7H11D6 bound to α2M-methylamine and α2M-trypsin but not to native α2M. The structure of α2M after conformational change resembled the letter "H." 7H11D6 epitopes were identified near the apices of the four arms in the α2M "H" structure. 7H11D6 that was adducted to colloidal gold (7HAu) retained the specificity of the free antibody (binding to α2M-trypsin but not to native α2M). α2M conformational change intermediates prepared by sequential reaction with a protein crosslinker and trypsin also bound 7HAu. These results suggest that a complete α2M conformational change is not necessary for 7H11D6 epitope exposure and may not be required for receptor recognition. 7HAu was used to isolate a preparation consisting primarily of binary α2M-trypsin (1 mole trypsin per mole α2M instead of 2). Structures resembling the letter "H" were most common; however, each field showed some atypical molecules with arms that were compacted instead of thin and elongated. These incompletely transformed structures were similar to the α2M conformational intermediates described previously (S. L. Gonias and N. L. Figler (1989) J. Biol. Chem. 264, 9565-9570). We propose that lateral arm extension is a critical step in α2M conformational change. Failure of lateral arm extension is probably a common property of different α2M conformational intermediates. © 1991.