ROLE OF AMINO-ACID POSITION-70 IN THE BINDING-AFFINITY OF P50.1 AND P58.1 RECEPTORS FOR HLA-CW4 MOLECULES

In an attempt to identify the amino acid position(s) of the HLA-C-spec ific p58.1/p50.1 natural killer cell receptors that determine the bind ing affinity for their ligand, we used soluble fusion proteins formed by the ectodomain of either receptor and the Fc portion of human IgG1. We show that the soluble p50.1 (activating) receptor binds weakly to 221-Cw4 transfectants. In contrast, the soluble p58.1 (inhibitory) rec eptor binds with high affinity. A single amino acid mutation at positi on 70, obtained by site-directed mutagenesis, was found to affect the binding affinity of both the p50.1 and the p58.1 receptors. Thus, subs titution in p50.1 of lysine 70 by threonine (typical of the inhibitory p58.1 molecule) resulted in a dramatic increase in binding affinity, comparable to that of the p58.1 molecule. On the other hand, substitut ion of threonine 70 by lysine in p58.1 almost abolished binding to 221 -Cw4 cells. Our present data indicate that a single amino acid differe nce greatly influences the p58.1/p50.1 affinity for their HLA-C ligand and suggests a possible role of position 70 as a contact site in the natural killer cell receptor/major histocompatibility complex class I interaction.