Structural and biophysical insights into T cell receptor recognition of lengthy antigens presented by major histocompatibility complex class I molecules

Abstract

Cytotoxic T cells represent an important line of defence in the vertebrate immune system to combat against intracellular abnormality such as viral and bacterial infections, as well as cellular transformation. In order to perform these functions effectively, cytotoxic T cells express αβ T cell receptors (αβ TCRs) on their cell surface, which allow them to specifically engage and differentiate self, altered self and foreign antigens on the surface of targeted cells. Intriguingly, such recognition is genetically restricted to antigen presentation by the host Major Histocompatibility Complex class I (MHC-I) molecules, which typically bind to short peptide fragments between 8 to 10 amino acids in length. Longer antigens on the other hand, have also been shown to represent potential targets for cytotoxic T cells, although it is not fully understood how TCRs can accommodate these peptide-MHC-I landscapes. Contrasting TCR specificity, TCRs also simultaneously exhibit remarkable ability to cross-react onto different targets. This binding degeneracy forms an essential part of the protective immunity, and allows TCRs to effectively recognise a diverse range of antigens that is presented to the host. Understanding the dual specificity of TCR (specificity versus degeneracy) is important, as it not only plays central roles in protective immunity but also contributes to clinical manifestation such as graft rejection and graft-versus-host diseases during organ transplantation. Using X-ray crystallography and Surface Plasmon Resonance (SPR) approaches as main techniques, my thesis set out to explore the underlying basis of simultaneous TCR specificity and cross-reactivity in the context of lengthy antigens (>10 amino acids) derived from a ubiquitous human pathogen, Epstein-Barr Virus (EBV). These antigens, when bound to closely related Human Leukocyte Antigen (HLA, MHC in human) molecules, HLA-B*35:08 and HLA-B*35:01, exhibit a range of non-canonical structural features, either bulging away from the antigen-binding platform or displaying marked conformational mobility. The structures presented in this thesis demonstrated that, in response to a super-bulged and rigid peptide antigen, two distinct TCR binding mechanisms could be employed. Namely, via the use of differing gene usages, TCRs can either engage and focus structurally and energetically onto the bulged antigen or, opt to bypass such a prominent feature by adopting an extreme docking orientation. These differences in the structural footprints in turn, allow TCRs to “see” or “ignore” subtle variations on the MHC landscape, and subsequently determine whether the TCR is MHC-restricted or cross-reactive. In addition to these findings, I also investigated the mechanism utilized by a TCR to accommodate lengthy and mobile antigen. Here, TCR recognition occurred via induced-fit of the antigen itself, which allows the optimal co-recognition of peptide and HLA landscapes to be achieved. Subtle sequence variations, such as HLA polymorphism or viral variants for instance, can modulate the shape and dynamic of the MHC bound antigen, which indirectly fine-tune TCR recognition and the subsequent biological responses. Taken together, these discoveries have not only provided novel kinetic and structural insights into lengthy detection by cytotoxic T cells, but also contributed towards our current understanding into the multifaceted nature of T-cell mediated cellular immune responses.