Novel Immunodominant Peptide Presentation Strategy: a Featured HLA-A*2402-Restricted Cytotoxic T-Lymphocyte Epitope Stabilized by Intrachain Hydrogen Bonds from Severe Acute Respiratory Syndrome Coronavirus Nucleocapsid Protein

Abstract

Antigenic peptides recognized by virus-specific cytotoxic T lymphocytes (CTLs) are presented by major histocompatibility complex (MHC; or human leukocyte antigen [HLA] in humans) molecules, and the peptide selection and presentation strategy of the host has been studied to guide our understanding of cellular immunity and vaccine development. Here, a severe acute respiratory syndrome coronavirus (SARS-CoV) nucleocapsid (N) protein-derived CTL epitope, N1 (QFKDNVILL), restricted by HLA-A*2402 was identified by a series of in vitro studies, including a computer-assisted algorithm for prediction, stabilization of the peptide by co-refolding with HLA-A*2402 heavy chain and ␤ 2 -microglobulin (␤ 2 m), and T2-A24 cell binding. Consequently, the antigenicity of the peptide was confirmed by enzyme-linked immunospot (ELISPOT), proliferation assays, and HLA-peptide complex tetramer staining using peripheral blood mononuclear cells (PBMCs) from donors who had recovered from SARS donors. Furthermore, the crystal structure of HLA-A*2402 complexed with peptide N1 was determined, and the featured peptide was characterized with two unexpected intrachain hydrogen bonds which augment the central residues to bulge out of the binding groove. This may contribute to the T-cell receptor (TCR) interaction, showing a host immunodominant peptide presentation strategy. Meanwhile, a rapid and efficient strategy is presented for the determination of naturally presented CTL epitopes in the context of given HLA alleles of interest from long immunogenic overlapping peptides.In 2003, severe and acute respiratory syndrome (SARS), emerging from China, caused a global outbreak, affecting 29 countries, with over 8,000 human cases and greater than 800 deaths (5, 9, 24, 33, 37). Thanks to the unprecedented global collaboration coordinated by the WHO, SARS coronavirus (SARS-CoV), a novel member of Coronaviridae family, was rapidly confirmed to be the etiological agent for the SARS epidemic (36). Soon after the identification of the causative agent, SARS was controlled and then quickly announced to be conquered through international cooperation on epidemiological processes (9). However, the role that human immunity played in the clearance of SARS-CoV and whether the memory immunity will persist for the potential reemergence of SARS are not yet well understood.In viral infections, CD8 ϩ cytotoxic T lymphocytes (CTLs) are essential to the control of infectious disease. Virus-specific CD8 ϩ T cells recognize peptides which have 8 to 11 amino acids, in most cases presented by major histocompatibility complex (MHC) class I molecules. However, identification of virus-specific CD8 ϩ T-cell epitopes remains a complicated and time-consuming process. Various strategies have been developed to define CTL epitopes so far. One of the most common practices to determine immunodominant CTL epitopes on a large scale is based on screening and functional analysis of overlapping 15-to 20-mer peptides covering an entire viral proteome or a given set of immunogenic proteins (...

Crystal Structure of Cell Adhesion Molecule Nectin-2/CD112 and Its Binding to Immune Receptor DNAM-1/CD226

“…Recombinant proteins were expressed in BL21 E. coli cells as inclusion bodies and were subsequently refolded and purified, as described previously (52,53). Biotinylation of proteins in vitro was achieved by incubating the protein with the Bir A biotin protein ligase, D-biotin, and ATP (52).…”

Structural Basis of Diverse Peptide Accommodation by the Rhesus Macaque MHC Class I Molecule Mamu-B*17: Insights into Immune Protection from Simian Immunodeficiency Virus

“…The characteristics of peptide presentation by MHC I play a pivotal role in the antigenicity of an epitope (44)(45)(46)(47)(48)(49). Accordingly, we determined the structures of eight SIV mac 239-derived epitopes in complex with Mamu-B*17 to define the atomic basis for effective epitope presentation by this protective MHC I molecule.…”

“…7A). On the other hand, the unique charged Glua 45 , which is located at the bottom of pocket B, also plays an (Fig. 7B) .…”

“…For QW9 (orange), the side chain of Thr 2 is too short to insert into the bottom of pocket B, which needs a water molecule to mediate the interactions among Thr 2 , Glua 45 , and Alaa…”

“…For FW9 (salmon) and GW10 (cyan), the side chains of Gln 2 and His 3 hydrogen bond to Trya 9 and Glua 45 .…”