An amino acid motif in HLA-DRß1 distinguishes patients with uveitis in juvenile idiopathic arthritis

A.J.W. Haasnoot; M.W. Schilham; S.S.M. Kamphuis; P.C.E. Hissink Muller; A. Heiligenhaus; D. Foell; R.A. Ophoff; T.R.D.J. Radstake; A.I. Den Hollander; T.H.C.M. Reinards; S. Hiddingh; N. Schalij-Delfos; E.P.A.H. Hoppenreijs; M.A.J. van Rossum; C. Wouters; R.K. Saurenmann; N. Wulffraat; R. ten Cate; J.H. de Boer; S.L. Pulit; J.J.W. Kuiper

Next, we tested the predicted affinities of all 83,686 overlapping 15-mer peptides from the selected 147 proteins in netMHCIIpan3.1. that tentatively could be presented by HLA-DR for binding to representative four-digit alleles of HLA-DRB1 (*01:01, *03:01, *04:01, *07:01, *08:01, *09:01, *10:01, *11:01, *12:01, *13:01, *14:01, *15:01, and *16:01). These alleles account for 79% of HLA-DRβ1 alleles in the JIA cases. The affinity data was log-transformed to a value between 0 and 1 using: 1−log(IC 50 nM)/log(50,000). 38 To categorize HLA-DRB1 allotypes (alleles) with similar predicted binding preferences, we performed unsupervised hierarchical clustering. Heatmaps were created based on the Euclidean distance measure and the Ward's linkage method using the MetaboAnalyst server. 82 We computed the ratio of the average binding affinity of HLA-DRβ1 molecules that contain Serine-11 in the peptidebinding groove (*03:01, *08:01, *11:01, *12:01, *13:01, and *14:01) over the average binding affinity of HLA-DRβ1 proteins that have other amino acids at this position (*01:01, *04:01, *07:01, *09:01, *10:01, *15:01, and *16:01) as a measure for the overall difference in predicted binding affinity for each peptide. A frequency distribution of the ratio of the binding affinities was plotted for the entire set of peptides (including low and nonbinding peptides), potential ligands for DRβ1 with a binding affinity stronger than half maximal inhibitory concentration (IC 50 ) of 500 nM, or peptides with high binding affinity for Serine-11 encoding alleles(IC 50 <50nM). correspond to presence of the given amino acid residue(s). Presence of serine (Ser) or aspartic acid (Asp) at position 11 in HLA-DRB1 (imputation info score = 1.11) was the top hit after initial association testing. Conditioning on either aspartic acid or serine revealed that the amino acids at positions 10-13, all well-imputed (imputation info = 1.08) and in perfect linkage disequilibrium, explain the bulk of the initial signal. Association statistics for tyrosine (position 10), threonine (position 12) and serine or glycine (position 13) are identical to the association statistics reported for serine (position 11) due to linkage disequilibrium. . The molecule is positioned to provide a view from the top of the peptide-binding groove. The beta-chain (DRß) is highlighted in orange. Amino acid serine at position 11 (red) is located in the bottom center of the peptide-binding groove of HLA-DRß1. Adjacent amino acid tyrosine at position 10 (green) and threonine at position 12 (blue) and serine at position 11 are displayed as spheres. 3D structure was produced using UCSF Chimera. 83