Before exit from the endoplasmic reticulum (ER), MHC class I molecules transiently associate with the transporter associated with antigen processing (TAP1/TAP2) in an interaction that is bridged by tapasin. TAP1 and TAP2 belong to the ATP‐binding cassette (ABC) transporter family, and are necessary and sufficient for peptide translocation across the ER membrane during loading of MHC class I molecules. Most ABC transporters comprise a transmembrane region with six membrane‐spanning helices. TAP1 and TAP2, however, contain additional N‐terminal sequences whose functions may be linked to interactions with tapasin and MHC class I molecules. Upon expression and purification of human TAP1/TAP2 complexes from insect cells, proteolytic fragments were identified that result from cleavage at residues 131 and 88 of TAP1 and TAP2, respectively. N‐Terminally truncated TAP variants lacking these segments retained the ability to bind peptide and nucleotide substrates at a level comparable to that of wild‐type TAP. The truncated constructs were also capable of peptide translocation in vitro, although with reduced efficiency. In an insect cell‐based assay that reconstituted the class I loading pathway, the truncated TAP variants promoted HLA‐B*2705 processing to similar levels as wild‐type TAP. However, correlating with the observed reduction in tapasin binding, the tapasin‐mediated increase in processing of HLA‐B*2705 and HLA‐B*4402 was lower for the truncated TAP constructs relative to the wild type. Together, these studies indicate that N‐terminal domains of TAP1 and TAP2 are important for tapasin binding and for optimal peptide loading onto MHC class I molecules.