MUTANTS OF HUMAN BETA-2-MICROGLOBULIN MAP AN IMMUNODOMINANT EPITOPE WITHIN THE 3-STRANDED BETA-PLEATED SHEET

Genetically engineered structural variants of human beta 2-microglobul in (beta 2m) were produced by sequence exchange with mouse beta 2m for the purpose of examining species-specific antigenic determinant expre ssion. For aggregate mapping, mouse and human beta 2m, which differ by 30% in their primary sequence of 99 amino acids, were prepared as chi meric (human X mouse) molecules and expressed in the FO-1 beta 2m-null human melanoma cell line. A chimera containing residues 1-69 from hum an beta 2m (and residues 70-99 from mouse beta 2m) induced expression of the epitopes defined by the anti-beta 2m monoclonal antibodies (mAb ) BBM.1, NAME-1, and L368; the reverse chimera did not, although HLA c lass I heavy chain was evident on the cell surface as determined with the TP25.99 mAb, For fine dissection of the epitopes defined by these mAbs, site-directed mutants of beta 2m were prepared by replacement of individual amino acids in human beta 2m with the dimorphic residue fr om mouse beta 2m. Substitutions were made at each divergent residue be tween positions 1 and 66 and, as controls for COOH-terminal modificati on, a series of residues between positions 75 and 94. Replacement of a mino acids 38, 44, and 45, but not 16 other dimorphic residues in the linear stretch from residue 1 to residue 66, resulted in the loss of, or gross reduction in, binding by mAbs BBM.1 and NAMB-1. A reduction i n binding was also observed for mAb L368. These data provide strong ev idence that the antigenic epitopes defined by these mAb map to a regio n including S3 and its adjacent intra-beta-strand turn of the three-st randed beta-pleated sheet of beta 2m. The mapping of these epitopes is consistent with their accessibility in the assembled major histocompa tibility complex class I molecule and indicates that the region from a mino acid 38 to 45 is an important structural feature in the ''foreign ness'' of human and mouse beta 2m.