ONE DISULFIDE BOND IN FRONT OF THE 2ND HEAVY-CHAIN CONSTANT-REGION ISNECESSARY AND SUFFICIENT FOR EFFECTOR FUNCTIONS OF HUMAN IGG3 WITHOUTA GENETIC HINGE

We have created four IgG3 mutants without a normal hinge region: (i) m 0 without a genetic hinge; (ii) m0/C131S, where Cys-131 in m0 was muta ted to Ser; (iii) m0/231C232 (formerly HM-1), where a Cys residue was inserted in m0 between Ala-231 and Pro-232; (iv) m0/C131S/231C232, whi ch is a hybrid of m0/231C232 and m0/C131S. The wild-type IgG3 and all mutants bind 5-iodo-4-hydroxy-3 -nitrophenacetyl groups. The wild type and mutants, m15 (with 15 aa in the hinge), m0/231C232, and m0/C131S/ 231C232, were all positive for complement-mediated lysis, antibody-dep endent cellular cytotoxicity mediated by peripheral blood leukocytes, and phagocytosis by U937. m0/C131S/231C232 was only weakly positive an d sometimes negative for respiratory burst activity mediated by periph eral blood neutrophils (polymorphonuclear leukocytes), whereas m15, m0 /231C232, and wild-type IgG3 were strongly positive. The m0 and m0/C13 1S mutants were mainly negative for complement-mediated lysis, antibod y-dependent cell-mediated cytotoxicity, and phagocytosis by U937 and p olymorphonuclear leukocytes. The results indicate that a hinge spacer region is not necessary, but the correct alignment of the two second h eavy chain constant regions in the IgG3 molecule by a minimum of one d isulfide bond is necessary and sufficient effector functions.