Class I HLA molecules “present” short peptide antigens to NK and T cell receptors to convert intracellular information into an extracellular immune response. These molecules are encoded by thousands of different alleles, each one adept in presenting distinct peptides. Differences in peptide presentation are an important factor in determining which alleles are best at alerting lymphocytes to the presence of a viral infection. While in silico methods exist to predict which peptides a particular HLA molecule binds, none can accurately predict the likelihood of presentation on the cell surface, nor the three-dimensional structure of a particular peptide-HLA complex. Thus, in vitro molecular and cellular experiments are essential to understand the properties of uncharacterized HLA molecules. In this thesis, I experimentally studied the antigen-presenting capabilities of two class I HLA molecules: the rare and archaic classical molecule, HLA-B*73:01 and the unusual and highly conserved non-classical molecule, HLA-F. The investigation of HLA-B*73:01 consists of a high resolution description of the peptides it presents (referred to as its peptidome) and the structural mechanisms that dictate how it presents these peptides. For HLA-F, I pursued two separate but overlapping lines of inquiry: understanding the different protein forms of HLA-F and whether they can both be detected on the cell surface; and how HLA-F is regulated in endometrial stromal cells in the immune privileged context of human pregnancy. While the study of HLA-B*73:01 highlights how unique it is relative to other class I molecules, the reappraisal of cell surface expressed HLA-F actually suggests that it is more like other class I molecules than previously thought.