The mouse NK inhibitory Ly-49A receptor specifically interacts with a peptide-induced conformational determinant on its MHC class I ligand, H- 2D(d). In addition, it binds the polysaccharide fucoidan, consistent with its C-type lectin homology and the hypothesis that Ly-49A interacts with carbohydrates on D(d). Herein, however, we demonstrate that Ly-49A recognizes D(d) mutants lacking N-glycosylation. Fucoidan competes for binding with anti-Ly-49A antibodies that inhibit Ly-49A-D(d) interaction, and blocks apparent Ly-49A binding to unglycosylated D(d). We confirm that Ly-49A recognizes the α1 and amino-terminal α2 domains of D(d) by analysis of recombinant H-2K(d)-H-2D(d) molecules. These studies indicate that Ly-49A recognizes carbohydrate-independent epitope(s) on D(d) and suggest that Ly- 49A has two distinct ligands, carbohydrate and MHC class I.