NETRAD_HLA
Description
Hypothesis: anti-HLA antibodies are NOT generated by exposure to mismatched HLA antigens, but are generated from viral coinfection (likely superinfection) with EBV + a distinct virus. Exosomes/viral components including viral attachment envelope proteins that exhibit neoepitopes are released under metabolic stress (as a result of SUMOylation and deglycosylation). The NETRAD algorithm is thoroughly described in "The Origins of Allograft Rejection" (pending publication in Human Immunology, Elsevier). During antigen presentation in regional lymph nodes, the germinal center light zone is more susceptible to oxygen deprivation and stress, facilitating slower antigen presentation of immune complexes via the neonatal Fc receptor (FcRn). Upregulation of L-lysozyme, among other lysosomal enzymes, enables the digestion and loading of deglycosylated peptides onto HLA to facilitate the recognition of neoepitopes. HLA-homologous peptide motifs were generated by the Smith-Waterman algorithm. This data consists of an MS Excel spreadsheet and MS Word document with accumulated peptide motifs demonstrating homologous amino acid sequence motifs between HLA and distinct virus pairs. Combinations of 6 peptides (including N-glycan signatures) can be mapped to HLA-specific epitopes.
Files
Steps to reproduce
See NETRAD algorithm MS Word Doc (attached). We acquired HLA or viral protein sequences from IMGT-HLA or NCBI. In silico N-lysozyme-digested viral peptide libraries were created utilizing EXPASY PeptideMass [20]. Viral peptides were assessed for HLA homology with the Smith-Waterman algorithm. Gapped viral peptide sequences, generated according to section 2.1, were superimposed upon correlate HLA crystal structures at the location of sequence homology and scored using an ectodomain-relative peptide (ERP) locating system. By utilizing the PDB database, pHLA3D and YASARA software, viral peptides were evaluated for antibody access to HLA- homologous sequences located on the correlate HLA protein. Peptide locations were scored as follows: 1= membrane-distal α-helices; 2= β-pleat membrane- distal side chains; 3= β-pleat membrane-proximal undersides; 4= stalk region, located between the β-pleat and membrane proximity; 5= membrane abutment; 0= cryptic locations lacking antibody-accessible residues (i.e., peptide cleft, buried or transmembrane residues). ERP scoring locations appear in Figure 1. ERP scores of “1-2” are positioned for optimal antibody-binding access in the extracellular space. ERP scores between “3-5” or “0” are considered cryptic locations, consistent with having been reported to lack clinical relevance [25]. Peptides were sorted by ERP score and amino acid sequence position of the mature HLA protein. We mapped combinations of viral peptides onto crystal structures and determined whether a clinically relevant HLA-specific epitope could be delineated to encompass the functional eplet, and if so, the minimum number of peptides necessary to do this. Importantly, HLA-epitopes derived from viral peptide combinations were determined to be clinically relevant only when an epitope was comprised of at least seven antibody- accessible amino acids, included the functional eplet to convey antibody specificity, and was encompassed within an established 15-angstrom radius. Additionally, HLA epitopes delineated by viral peptides must be unimpeded by the naturally presented peptide held within the HLA cleft. This ensured that HLA cleft-presented peptides would not subject the epitope to continuous amino acid residue variation. PTM sequons, specifically for Small Ubiquitin-like Molecules (SUMO) and N-glycans that might influence the generation of neoepitopes under oxygen-deprived conditions, were identified independently for each viral envelope protein (Figure 2) and each viral peptide sequence, respectively (Tables 2-3).