Peptide editing and MHC binding mechanisms of Tapasin and TAP binding protein related, TAPBPR

Abstract

Abstract Cell surface major histocompatibility complex class I (MHC-I) molecules play a crucial role in immunity to microbes and tumors by presenting intracellular peptides for recognition by CD8+ T cells. Since the repertoire of bound peptides is crucial to MHC-I stability as well as to T cell and NK cell recognition, the molecular mechanism by which peptides are selectively loaded onto MHC-I molecules is of considerable interest. A key role is played by the well-characterized chaperone tapasin as indicated by reduced cell surface MHC-I levels and altered peptide repertoires in tapasin deficient cell lines and mice. Despite extensive studies of the functional role of tapasin in shaping peptide repertoires, direct structural information on its interaction with MHC-I is lacking. Following the recent biochemical and crystallographic studies of the interaction of the tapasin homolog, TAPBPR, with MHC-I, we have engineered soluble versions of tapasin for comparison with TAPBPR. Using surface plasmon resonance assays of purified, untethered proteins, we observe that tapasin binds directly to an MHC-I molecule loaded with truncated peptides but with lower apparent affinity than TAPBPR. Similarly, using fluorescence polarization, we find that tapasin facilitates peptide exchange onto MHC-I with a lower efficiency than TAPBPR. These experiments permit direct comparisons of tapasin/MHC-I and TAPBPR/MHC-I interactions and help elucidate the functional relationship of these two distinct chaperones in shaping the MHC-I peptide repertoire.