Analysis Of The Monomer And Complex Crystal Structures Of Bovine CD8αα/BoLA-I And The Function Of Presenting Antigenic Peptides

The main histocompatibility complex class1molecule presents viral epitopes (pMHC1) and is recognized by T cell receptor (TCR) and the CD8co-receptor molecules, this formation of an immune synapse results in the proliferation of cytotoxic lymphocyte (CTL), induction cellular immune responses against specific pathogens, lyses and eventual clearance of the target cells. About the researching of MHC I molecules, compared with human and mouse, there is a few research in structural studies about bovine leukocyte antigen I (BoLA-I). These years, viral diseases and other infections have been caused by zoonosis of cattle, such as foot-and-mouth disease virus (FMDV) which brought great economic losses and post a threat to human health,. So far, there are a few studies of the bovine cellular immune response which mediated by viruses, especially the mechanism of BoLA-I presenting viral antigen peptides and CD8as the auxiliary receptor interaction with MHC I molecules in CTL immune response is not clear. Therefore, father studies of the refined crystal structures of the monomer and complex of bovine MHC I (BoLA-I) and CD8(bCD8) are essential in order to clarify the mechanism of immune response of CTL and overcome bovine-origin Zoonotic diseases. Here, with the methods of molecular cloning and structural biology, the crystal structure of BoLA (BoLA-2*02201) complexed with peptide derived from the FMDV virus was solved. Furthermore, we solved the bCD8aa homodimer structure. Finally, CD8aa/BoLA-I crystal structure was also determined. We first acquired immunological data of bovine CTL immune response.The overall structure of bovine BoLA-I is similar with the structures of human and mouse MHC I which have been determined, but has the characteristic of particular species or genera. Compared the structure with other two BoLA-I molecules, there are some prominent structural features of BoLA-2*02201. It has a large "open" A pocket, negatively charged B pocket and deeply and the largest volume of the F pocket. The peptide which can be presented by BoLA-2*02201has such characteristics:1) the anchor residues of P1can be larger conformation amino acids;2) P2positively charged with long side chains of amino acid,3) the anchor residues of P9position should be with a larger aromatic rings. According to the results of structure analysis and peptide binding, we put forward the peptide binding motif of the BoLA-2*02201molecule.After modification the gene of CD8aa, it can be expressed and folded as bovine CD8aa homodimer, and its structural was crystallized and determined. According to the residues of CD8aa which interact with MHC I in human and mouse CD8aa/pMHC I complexes, the three R6, N102and S103amino acids are relatively conservative in bovine CD8aa molecule, these may be the residues which mediated the interaction between bovine CD8αα and MHC I molecules.Finally, the results of bovine CD8aa/BoLA-2*02201complex are as follows:1) Bovine CD8aa of CD8aa/BoLA-2*02201complex cannot change the conformation of the peptide;2) After contacted with bovine CD8αα, the main change of BoLA-2*02201is offset direction of α3CD loop;3) The change of bovine CD8αα is mainly about the offset direction of CDR like loops.4) Compared with human and mouse CD8aa/MHC I complexes, the bovine CD8aa/BoLA-2*02201complex has least contaction area and minimal interactions. Determinately, the interaction between bovine CD8αα and BoLA-I is in antigen-antibody contact manner.In brief, we solved the crystal structure of BoLA-I (BoLA-2*02201) and illustrate the structure basis of BoLA-I presenting peptides, and further screened and identified some potential CTL epitopes of FMDV-VP1. In addition, the crystal structure of bCD8aa was also determined, and carified the structure characteristics of bCD8aa. Analysis of the crystal structure of bovine CD8aa/BoLA-2*02201complex, we found the species characteristics between bovine CD8aa and BoLA-2*02201interaction, and it has least contaction area and minimal interactions.In conclusion, these results might promote the development of the structural immunology of CTL immune response in bovine cellular immunology.