Killer Immunoglobulin-like receptors (KIRs) are a family of receptors with either activating or inhibitory signalling potential that are integral to triggering Natural Killer Cell mediated cytolysis. The ligands for KIRs are peptide-loaded human leukocyte antigen (pHLA) class I molecules. It is known that allelic variation and polymorphism within the KIR family affects pHLA binding specificity and is linked to the control of viral replication and the treatment outcome of certain malignancies. However, the molecular details linking allotypic differences with disease progression have proven elusive. We have utilised structural biology and extensive mutagenesis to provide a framework for understanding the intricate interplay between peptide variability, KIR polymorphism and HLA polymorphism in determining KIR specificity. By mapping the interaction site we detail a common basis for KIR-pHLA recognition. Furthermore, we show that residues beyond the interface are critical to determining allotype specificity.