THE EFFECT OF CYSTEINE MODIFICATION AND PROTEINASES ON THE MAJOR ANTIGENS (D, C, C, E AND E) OF THE RH BLOOD-GROUP SYSTEM

We have confirmed and extended previous observations showing that the (Rh) D antigen of erythrocyte membranes is destroyed by various reagen ts that modify cysteine (Cys) residues (Res.) and by trypsin as well a s chymotrypsin, using thirty examples of monoclonal or polyclonal anti -D in heamglutination inhibition assays. We have also shown that most C, c, E, e and BS58 epitopes are inactivated or weakened by most Cys r eagents and by these proteinases, using monoclonal and polyclonal anti bodies. Inactivation by 5,5-dithiobis-(2-nitrobenzoic acid) was always fully reversible after subsequent dithioerythritol treatment. The ess ential Cys Res. appear to be buried in the membrane in view of the ina bility of some reagents to inactivate (iodoacetamide, iodoacetic acid) or reactivate (reduced glutathione) the antigens. Data obtained with N-ethylmaleimide indicate that inactivation of the C and c antigens is , at least in part, attributable to (a) Cys Res. that is (are) differe nt from that (those) involved in the E and e antigens. Data obtained w ith the Cys reagents and the proteinases suggest that more than one pe ptide loop of the Rh proteins is involved in the major Rh antigens.