Visualization of inhibitory Ly49 receptor specificity with soluble major histocompatibility complex class I tetramers

Murine natural killer (NK) cells are inhibited from killing their targets b y the interaction between inhibitory, C-type lectin like Ly49 receptors and major histocompatibility complex (MHC) class I molecules. The receptors ha ve overlapping specificity, and it has been difficult to analyze specific a spects of the interaction between different Ly49 receptors and their respec tive ligands. We have addressed this problem using tetramers of bacterially expressed, non-glycosylated, MHC class I molecules refolded with different peptides. Our results indicate that this technology is useful for analysis of Ly49 receptor specificity as well as for monitoring of NK cell subsets, with the following major conclusions emerging from this study: (1) tetrame rs of H-2D(d) bound the Ly49A receptor; the MHC associated glycan, previous ly suggested to be involved in recognition by this receptor, is thus not re quired for Ly49A receptor binding; (2) in support and extension of a recent report indicating peptide selectivity in the recognition of H-2K(b) by Ly4 9C(+) cells, H-2K(b) tetramer binding to Ly49C receptors was strongly influ enced by the peptide presented by the MHC class I molecule; (3) tetramer bi nding allowed visualization of interactions that have not previously been d etected in functional studies, such as the recognition of H-2D(b) by Ly49A and Ly49C.