Peptide-conformed β₂m-free class I heavy chains are intermediates in generation of soluble HLA by the membrane-bound metalloproteinase

Molecular mechanisms of soluble HLA-release by a membrane-bound metalloproteinase (MPase) are not defined. We have investigated the possibility that certain β₂-microglobulin (β₂m)-free heavy chains (HC) retain peptide-induced conformations before and after the cleavage by using mutant HLA-A2.242K HC with reduced affinity for β₂m. We show that dissociation of HC/β₂m complexes on the surface of C1R lymphoblastoid cells generates both conformed and non-conformed β₂m-free HC recognized by conformation-dependent antibodies. Conformed HC, having bound the HLA-A2- specific peptide HTLV-1 tax 11-19, can retain their proper conformations after dissociation of β₂m. Further, conformed and non-conformed surface β₂m-free HC are cleaved by the MPase, and some released HC preserve their conformations. Exogenous β₂m binds only to conformed HC, and protects them from cleavage as effectively as the MPase inhibitor BB-2116. We propose that soluble HLA-release requires generation of peptide-conformed β₂m-free HC intermediates on the cell surface, which are then cleaved by the MPase and in solution may reassociate with β₂m. Given the role of soluble HLA in the indirect allorecognition, the activity of this MPase may be important in transplant rejection.