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. 2017 May 8;12(5):e0177140.
doi: 10.1371/journal.pone.0177140. eCollection 2017.

Citrullination Only Infrequently Impacts Peptide Binding to HLA Class II MHC

Free PMC article

Citrullination Only Infrequently Impacts Peptide Binding to HLA Class II MHC

John Sidney et al. PLoS One. .
Free PMC article


It has been hypothesized that HLA class II alleles associated with rheumatoid arthritis (RA) preferentially present self-antigens altered by post-translational modification, such as citrullination. To understand the role of citrullination we tested four RA-associated citrullinated epitopes and their corresponding wild-type version for binding to 28 common HLA class II. Binding patterns were variable, and no consistent impact of citrullination was identified. Indeed, in one case citrullination significantly increased binding compared to the WT peptide, in another citrullination was associated with a reduction in promiscuity by 40%. For a more comprehensive analysis, we tested over 200 citrullinated peptides derived from vimentin and collagen II for their capacity to bind the RA-associated shared epitope alleles DRB1*01:01 and DRB1*04:01. The overall effect of citrullination on binding was found to be relatively minor, and only rarely associated with 3-fold increases or decreases in affinity. Previous studies have suggested that citrullination of MHC anchor residues, in particular P4, is associated with generation of novel RA-associated epitopes. However, analysis of the predicted MHC-binding cores of all peptides tested found that in modified peptides with increased binding affinity the citrullinated residue was predicted to occupy an anchor position in only a minority of cases. Finally, we also show that identification of citrullinated peptide binders could be facilitated by using the NetMHCIIpan 3.1 algorithm, representing citrullination as a wildcard. Our studies identify a total of 117 citrullinated peptides that bound RA-associated alleles with an affinity of 1000 nM or better.

Conflict of interest statement

Competing Interests: This commercial affiliation does not alter our adherence to all PLOS ONE policies on sharing data and materials.


Fig 1
Fig 1. Comparison of binding affinity of wild-type and citrullinated versions of vimentin peptides.
Each data point indicates the DRB1*01:01 (left panel) or DRB1*04:01 (right panel) binding capacity of WT vimentin peptides with the corresponding citrullinated version. Effects greater or less than 3-fold are demarcated by the diagonal dashed red lines and highlighted by red fill. Points to the lower right indicate instances where the citrullinated peptide binds with higher affinity that the WT peptide, and vice versa.
Fig 2
Fig 2. Prediction of the DRB1*01:01 and DRB1*04:01 binding capacity of citrullinated peptides.
The DRB1*01:01 (left panel) and DRB1*04:01 (right panel) binding capacity of citrullinated peptides were predicted using NetMHCIIpan version 3.1 predictions by substituting the citrullinated residues with the wildcard “X”. Trend lines are show in red.
Fig 3
Fig 3. Increases in binding capacity due to citrullination is associated with modification at both anchor and non-anchor positions.
The number of instances of increased DRB1*01:01 (left panel) or DRB1*04:01 (right panel) binding associated with citrullination of arginine at specific peptide positions relative to predicted core region frames is shown. Anchor positions are highlighted with red bars and non-anchor by blue bars. Light filled bars show the total number of increases attributed to anchor (white filled red bars) and non-anchor (blue hatched bars) positions. Tabulations are shown for two different approaches to defining the core residues, as described in the text.

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Grant support

These collaborative studies have been supported by funding from Janssen Research and Development. Janssen Research and Development provided financial support in the form of authors' salaries (SB, KD, NR, TR, MZ), but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section. Additional support has been provided from the NIH-NIAID Immune Epitope Database and Analysis Project [contract HHSN272201200010C] (AS).

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