The HLA‐E class Ib molecule constitutes a major ligand for the lectin‐like CD94/NKG2 natural killer (NK) cell receptors. Specific HLA class I leader sequence‐derived nonapeptides bind to endogenous HLA‐E molecules in the HLA‐defective cell line 721.221, inducing HLA‐E surface expression, and promote CD94/NKG2A‐mediated recognition. We compared the ability of NK clones which expressed either inhibitory or activating CD94/NKG2 receptors to recognize HLA‐E molecules on the surface of 721.221 cells loaded with a panel of synthetic nonamers derived from the leader sequences of most HLA class I molecules. Our results support the notion that the primary structure of the HLA‐E‐bound peptides influences CD94/NKG2‐mediated recognition, beyond their ability to stabilize surface HLA‐E. Further, CD94/NKG2A⁺ NK clones appeared more sensitive to the interaction with most HLA‐E‐peptide complexes than did effector cells expressing the activating CD94/NKG2C receptor. However, a significant exception to this pattern was HLA‐E loaded with the HLA‐G‐derived nonamer. This complex triggered cytotoxicity very efficiently over a wide range of peptide concentrations, suggesting that the HLA‐E/G‐nonamer complex interacts with the CD94/NKG2 triggering receptor with a significantly higher affinity. These results raise the possibility that CD94/NKG2‐mediated recognition of HLA‐E expressed on extravillous cytotrophoblasts plays an important role in maternal‐fetal cellular interactions.