A MUTANT HUMAN BETA(2)-MICROGLOBULIN CAN BE USED TO GENERATE DIVERSE MULTIMERIC CLASS-I PEPTIDE COMPLEXES AS SPECIFIC PROBES FOR T-CELL RECEPTORS

Antigen-specific receptors (TCR) on CD8 T lymphocytes form relatively short-lived complexes with their natural ligands: peptides in associat ion with major histocompatibility complex (MHC) class I molecules, whi ch consist of a polymorphic heavy chain and a conserved light chain, b eta(2)-microglobulin (beta(2)-M). To produce soluble MHC-peptide compl exes in a form that would bind more stably and could be used to identi fy, count, and isolate CD8 T cells having the appropriate TCR, we prep ared multimeric MHC-peptide complexes, Our work builds on the assembly of recombinant MHC class I peptide complexes using a mutant human bet a(2)-M chain (Tyr 67 > Cys) which can form stable heterodimers with di verse MHC heavy chains. With biotin added to the SH group, the assembl ed MHC-peptide monomers formed multimers with avidin linked to a fluor ochrome. The specific reactivity of the multimeric reagents with human and mouse cytotoxic T cells (CTL) is described, The present approach permits the production of class I multimers, without the necessity of genetic engineering each heavy chain, a significant advantage in view of the enormous polymorphism of MHC heavy chains, Because human beta(2 )-M forms stable heterodimers with diverse class I heavy chains from v arious species (human and non human primates, mouse, etc.), this proce dure is a general method for producing multimers of MHC-peptide comple xes as T cell receptor-specific probes. (C) 1998 Elsevier Science B.V. All rights reserved.