Peptide length significantly influences in vitro affinity for MHC class II molecules

Immunome Research BioMed Central Research Open Access Peptide length significantly influences in vitro affinity for MHC class II molecules Cathal O'Brien1, Darren R Flower2 and Conleth Feighery*1 Address: 1Department of Immunology and Institute of Molecular Medicine, St James's Hospital and Trinity College Dublin, Ireland and 2The Jenner Institute, University of Oxford, Compton, Berkshire, RG20 7NN, UK Email: Cathal O'Brien - ; Darren R Flower - ; Conleth Feighery* - * Corresponding author Published: 26 November 2008 Immunome Research 2008, 4:6 doi:10.1186/1745-7580-4-6 Received: 26 June 2008 Accepted: 26 November 2008 This article is available from: http://www.immunome-research.com/content/4/1/6 © 2008 O'Brien et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: Class II Major Histocompatibility Complex (MHC) molecules have an open-ended binding groove which can accommodate peptides of varying lengths. Several studies have demonstrated that peptide flanking residues (PFRs) which lie outside the core binding groove can influence peptide binding and T cell recognition. By using data from the AntiJen database we were able to characterise systematically the influence of PFRs on peptide affinity for MHC class II molecules. Results: By analysing 1279 peptide elongation events covering 19 distinct HLA alleles it was observed that, in general, peptide elongation resulted in increased MHC class II molecule affinity. It was also possible to determine an optimal peptide length for MHC class II affinity of approximately 18–20 amino acids; elongation of peptides beyond this length resulted in a null or negative effect on affinity. Conclusion: The observed relationship between peptide length and MHC class II affinity has significant implications for the design of vaccines and the study of the epitopic basis of immunological disease. Background Classical major histocompatibility complex (MHC) molecules are divided into two groups: class I and class II [1]. Each contains large numbers of alleles, all of which bind antigen for cell surface presentation to T cells. MHC class I molecules comprise a single polymorphic chain coupled to a single conserved protein: β2 microglobulin. In contrast, MHC class II molecules comprise two polymorphic subunits: an α and a β chain. A notable difference between the classes is the open-ended peptide-binding groove of MHC class II molecules compared to the closed binding site of MHC class I. This feature, which has been observed in many X-ray crystallographic structures, has been used to explain the observed differences in peptide length accommodated by the two classes [2-4]. Class I molecules typically accommodate peptides of eight to ten residues, although instances of longer peptide binding have now been reported [5]. In contrast, MHC class II molecules can accommodate much longer peptides. Stern et al. investigated an influenza-derived peptide complexed with HLA-DR1; they saw the peptide bound in an extended conformation with five binding pockets, which engaged peptide side chains, while the flanking regions extended out of the groove [3]. Since MHC class II molecules have an open-ended bindPage 1 of 7 (page number not for citation purposes) Immunome Research 2008, 4:6 ing groove, they do not, in general, restrict the length of bound peptides. The length of peptides occupying the groove can vary considerably as a consequence of semistochastic proteolytic degradation [6]. Several studies have shown the influence of residues outside the main nonameric core on binding and subsequent T cell recognition. Residues outside the nonameric binding region but within an extended binding groove have been called peptide-flanking residues (PFRs) [7]. The importance of PFRs for T cell recognition has also been investigated. It is widely accepted that these flanking regions can contribute to T cell mediated pMHC recognition [8]. Arnold and colleagues showed that certain T cell responses were completely dependent on residues at peptide positions P-1 and P11. Similar findings were reported by Stepniak et al for immunostimulatory HLA-DQ2 binding peptides derived from gliadin and glutenin proteins [9]. The findings of both authors can be partly explained by the results of X-ray crystallographic studies of peptideMHC class II-TCR trimolecular complexes [10,11]. These reports indicate that the CDR3 regions of the TCR α and β chains tend to locate over the p5 residue in the binding groove allowing the TCR to interact with the PFRs. It has also been shown that the properties of residues in the PFRs can influence T cell recognition, with residues capable of forming salt-bridges or hydrogen bonding with the TCR being most favoured [8]. Much circumstantial if anecdotal evidence suggests that PFRs contribute strongly to peptide-MHC stability. They can provide an increased measurable affinity of peptide for the MHC binding groove, in particular the residue at position P-1 [12]. Nelson et al. reported that the effect of increased stability was greater for a peptide length increase of six amino acids compared to four. They also found the effect to differ depending on which terminus was elongated. Moreover, a previous study by Srinivasan et al showed increased affinity resulting from the peptide elongation of a single peptide [13]. How such peptide elongations might increase affinity remains unclear. Although it is theoretically possible PFRs interact with the MHC molecules outside the groove, this has yet to be shown experimentally, nor has it been seen in X-ray crystallographic structures. We have sought to elucidate this phenomenon further, by examining the relationship between peptide length and MHC class II affinity. We devised custom algorithms and applied them to data available in the AntiJen database [14] to firstly ascertain whether peptide length can impact on affinity of a peptide for an MHC class II molecule. This was assessed by searching the database for instances of peptide elongation and determining the reported effect on MHC class II affinity. Following this, we examined http://www.immunome-research.com/content/4/1/6 whether there was a demonstrable limit to the effect of peptide length on MHC class II affinity and subsequently determined whether any effects on affinity were peptide terminus specific. Our analysis indicated the positive impact of peptide length on affinity. We also identified an optimal length for peptide-MHC class II binding and demonstrated that the effects on peptide affinity are terminus independent. Results Most peptide elongation events result in enhanced affinity The relationship between peptide length and MHC class II affinity was examined using a da (...truncated)