| Major histocompatibility complex (MHC) class I-peptide complexes present at the cell surface enable cytotoxic T-lymphocytes to recognize and eliminate malignant and infected cells. The assembly of MHC class I-peptide complexes in the endoplasmic reticulum is orchestrated by multiple proteins collectively referred to as the peptide-loading complex. Within the peptide-loading complex, tapasin plays a crucial role in optimizing peptide selection, which results in efficient antigen presentation. A central issue remaining in the field of MHC class I biology is the molecular mechanism by which tapasin facilitates peptide editing. During peptide loading, MHC class I molecules are hypothesized to transition through an intermediate conformer before achieving a fully folded state. Therefore, I took a serologic approach to define a distinct, tapasinassociated MHC class I conformation, which likely represents intermediately folded MHC class I molecules. In other studies, I utilized specific tapasin mutants to reveal protein interactions within the peptide-loading complex, and to determine the significance of these interactions for tapasin function. The tapasin transmembrane/cytoplasmic region was found to be exceptionally important for the surface expression of murine MHC class I molecules. From these studies, I proposed a model whereby the tapasin transmembrane/cytoplasmic domains are important both for interactions with the transporter associated with antigen processing and with the MHC class I molecule itself. My investigations utilizing a tapasin mutant lacking an exposed loop within the tapasin C-terminal immunoglobulin-like domain emphasize that this region is an important component of the tapasin-MHC class I interface. Additionally, through molecular modeling, I have predicted specific interactions between tapasin and the MHC class I peptide-binding groove and alpha3 domain. In studies in which I characterized a tapasin cysteine mutant, a novel disulfide-linked complex involving tapasin, ERp57, and the MHC class I heavy chain was identified. Furthermore, in these studies tapasin C95 was found to be necessary for optimal MHC class I surface expression. Overall, this work sheds new light on tapasin's function in promoting MHC class I assembly and also provides novel insight into folding of the MHC class I peptide-binding groove. |
