SUBSTITUTION OF CLASS-II ALPHA-CHAIN POLYMORPHIC RESIDUES DEFINES LOCATION OF A(ALPHA)(K) SEROLOGIC EPITOPES AND ALTERS ASSOCIATION BETWEENALPHA-BETA AND II POLYPEPTIDES

Structure-function studies of the MHC class II alpha chain have been p erformed by constructing a panel of A(alpha)k CDNA genes with one or m ore d allele substitutions at each polymorphic residue of the alpha1 d omain. The altered genes (A(alpha)k) were transfected into a B lympho ma cell line (BKO), which is deficient in Aalpha mRNA but retains cons titutive wild-type A(beta)k mRNA expression. Cytofluorometric analysis of cell surface A(alpha)k A(beta)k molecules was used to map the pol ymorphic alpha chain residues comprising four serologic epitopes. A(al pha)k-reactive mAbs 1E9, 2A2, and 3F12 recognize an epitope that inclu des the polymorphic residue 44 of the A(alpha)k polypeptide, and the A (alpha)d-reactive antibody, K24-199, recognizes a conformationally det ermined epitope influenced by residues from all three polymorphic regi ons. In addition, we confirmed previous studies demonstrating that Ia. 19 and Ia.2 mAbs bind to epitopes adjacent to residue 75 in A(alpha)k. A cell surface negative expression variant also was identified in the panel of mutant cell lines and biochemically characterized. Substitut ion of six d allele polymorphic residues at positions 11, 14, 28, 69, 70, and 75 in the A(alpha)k polypeptide (T.EG A(alpha)k A(beta)k) res ults in an A(alpha)k polypeptide that associates with the A(alpha)k p olypeptide but does not associate with the Ii polypeptide. This defect in Ii - A(alpha)(k)A(alpha)k interaction is associated with a conform ational change in the alpha1beta1 domain that was identified by altere d reactivity of the T.EG complex with conformationally dependent anti- alpha and anti-beta mAbs. The bulk of the T.EG A(alpha)k. A(beta)k co mplexes contain high mannose oligosaccharides and, therefore, do not e xit the endoplasmic reticulum/cis-Golgi. A small fraction of the T.EG A(alpha)k A(beta)k, complexes are recognized by conformation dependen t antibodies and are competent to be transported at least to the media l Golgi.